title: Tag8: Monkey AC3, NP3, AC4

status: Public on Jun 07 2023

city: Suita

first: Noriyuki

last: Tsumaki

state: Select One

address: 2-2 Yamadaoka

department: Department of Biochemistry and Molecular Biology

country: Japan

institute: Osaka University, Graduate School of Medicine

data_processing:
TAS-Seq data cDNA reads were mapped to the reference genome (Macaca_fascicularis) using HISAT2-2.2.1(37) and the following parameters: -q -p 6 –rna-strandness F –very-sensitive –seed 656565 –reorder –omit-sec-seq –mm. For the HISAT2 index build, a corresponding ensembl gtf file was filtered to retain protein-coding RNA, long non-coding RNA, and T cell chain/immunoglobulin chain annotations according to the 10X Genomics’s method (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/advanced/references#mkgtf). Then, the cell barcode information of each read was added to the HISAT2-mapped BAM files, and associated gene annotations were assigned using featureCounts v2.0.2(38) and the following parameters: -T 2 -Q 0 -s 1 -t gene -g gene_name –primary -M -O –largestOverlap –fraction -R BAM. In the featureCounts analysis, a “gene” annotation was used to capture unspliced RNA information for the RNA velocity analysis, and primary annotations were kept.
Assembly: Macaca_fascicularis_6.0
Supplementary files format and content: tab-delimited text files include raw tag-count values for each sample
Supplementary files format and content: cell name description, each cell was named as follows: [experiment ID]:[hashtag ID]_[experiment ID]_[cell barcode].bam

organism: Macaca fascicularis

source_name: Nucleus pulposus (NP), Articular cartilage (AC)

extract_protocol:
The NP sample was gelatinous, and AC samples were minced into small pieces (2 mm) using a biopsy punch. Next, both samples were washed with preparation medium (RPMI-1640 [Nacalai Tesque] with 0.2% FBS [Gibco] and 10 mM HEPES) and transferred to a 6-well plate containing digestion medium (preparation medium with 0.2 mg/ml Liberase TM (Roche) and 2 kU/ml DNaseⅠ (Merck) within 2 h of sacrifice. The plate was then placed on a shaker and incubated for 3 h at 37 °C and 5% CO2. After dissociation, the samples were applied to a 70 μm cell strainer (BD Biosciences), followed by the next step of the cell barcoding process.   For cell surface labeling, we biotinylated cell surface proteins as described (M. Sugimoto, Y. Tada, S. Shichino, S. Koyamatsu, N. Tsumaki, K. Abe, Universal Surface Biotinylation: a simple, versatile and cost-effective sample multiplexing method for single-cell RNA-seq analysis, DNA Res. 29(3) (2022).). Cells were resuspended in 1 mL PBS supplemented with 1% FBS and 1 ng EZ-Link Sulfo-NHS-Biotin (Thermo Scientific) for 10 min at 4 °C, washed twice with PBS supplemented with 3% FBS, and then washed with Cell Staining Buffer (BioLegend). Next, the cells were stained with 0.6 μg/ml of Totalseq (A0951-A0954 [BioLegend]) for 20 min at 4°C for cellular barcoding, washed with Cell Staining Buffer, washed with PBS supplemented with 1% FBS, and resuspended in PBS supplemented with 10% FBS and 1 μM Sytox blue dead cell stain (Invitrogen) for 5 min at room temperature.  Barcoded live dissociated cells (PE-positive and Sytox blue-negative) were sorted and suspended in PBS supplemented with 20% FBS. The data was collected on a FACS Aria II flow cytometer (BD Bioscience) and analyzed with BD FACS Diva 9.0.1 (BD Biosciences).
Library preparation and the first processing of paired-end fastq files were performed according to the workflow in the previously reported TAS-seq method (Shichino S, Ueha S, Hashimoto S, Ogawa T, Aoki H, Wu B, et al. TAS-Seq is a robust and sensitive amplification method for bead-based scRNA-seq. Communications biology. 2022;5(1):602.). In brief, sorted cells were used to synthesize cDNA with BD Rhapsody Express Single-Cell Analysis System (BD Biosciences). cDNAs were amplified by the TAS-seq method at Immunogeneteqs Inc. (Noda city, Chiba, Japan) and sequenced with an Illumina Novaseq 6000 sequencer (Illumina, San Diego, CA, USA), using a Novaseq 6000 S4 reagent kit v1.0 or v1.5.

value: male

value: 3 year old

value: wild type


title: WTA8: Monkey AC3, NP3, AC4

type: SRA

library_source: transcriptomic single cell

molecule: polyA RNA

tag: tissue

value: Nucleus pulposus (NP), Articular cartilage (AC)

tag: Sex

tag: age

tag: genotype


title: mRNA-derived cDNA R1001_1-G

status: Public on Jun 16 2023

city: New York

first: Darwin

last: D'Souza

state: NY

address: 1470 Madison Avenue

department: ImmunoBio

country: USA

institute: Human Immune Monitoring Center

description:
mRNA-derived cDNA
polyA RNA
paired-end
Illumina NovaSeq 6000
5' mRNA library: read1 file contains cell barcode and UMI; read2 file contains transcript
Tumor, Normal, Lymph Node and PBMC

library_source: transcriptomic

data_processing:
Pre-processing of sequencing results to generate count matrices (gene expression, ADT and HTO barcode counts) was performed using the 10x genomics Cell Ranger pipeline where ADT and HTO sequences were concatenated and treated as Custom library with default settings (v3.0.2) and aligning to the hg19 1.2.0 genome build and Ensembl 82 annotations.
Further processing was done with Seurat v3.0.0 (cell and gene filtering, hashtag identification, clustering, differential gene expression analysis).
Assembly: hg19 1.2.0
Supplementary files format and content: Processed counts from CellRanger in an R object, data_SingleCellExperiment_object.Rda; Additional metadata in “sample_annots.csv” and “full_HCC_cluster_annotation.csv”

molecule: total RNA

organism: Homo sapiens

source_name: Liver

extract_protocol:
Human tumor lesions and non-involved adjacent tissues were digested as described in the material and method. Briefly, samples received mechanical and enzymatic digestion, filtration and the cell suspension was enriched in immune cells using a Percoll gradient, followed by a RBC lysis. Lymph nodes were digested mechanically and enzymatically followed by RBC lysis. Blood samples were processed by Ficoll followed by RBC lysis.
Library was prepared using Chromium Single Cell 3ʹ v2 protocol/3ʹ  protocol (10x Genomics). Cellular mRNA and antibody-derived oligos were reverse-transcribed and indexed with a shared cellular barcode. Indexed cDNAs were then pooled and amplified by PCR according to the Chromium Single Cell associated protocol (10x Genomics) and with specific primers amplifying the antibody-derived tags. SPRI bead size selection was performed in order to separate both the mRNA-derived cDNA (>300bp) and the antibody-derived tagged cDNAs (180bp). For the mRNA derived cDNA library preparation, we further proceeded according to the manufacturer’s instructions for Single Cell associated protocol (10x Genomics). For antibody-derived tagged library, we used the KAPA HiFI HotStart Library Amplification Kit (Roche) with the following primers and amplification program:
-CITE-seq library: amplification was performed at 95°C for 3’ followed by ten cycles at 95°C f¬¬or 20’’; 60°C for 30’’; 72°C for 20’’ and final elongation 72°C for 5’ with 5’-CAAGCAGAAGACGGCATACGAGATCGTGATGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA-3’ Small RNA RPI1 primer as i7 primer (Illumina).
-HASH-seq library: amplification was performed at 95°C for 3’ followed by ten cycles 95°C for 20’’; 64°C for 30’’; 72°C for 20’’ and final elongation 72°C for 5’ with 5’-CAAGCAGAAGACGGCATACGAGATCGAGTAATGTGACTGGAGTTCAGACGTGTGC-3’ TruSeq D701_s primer as i7 primer (Illumina).
The following 5’- AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTC-3’ SI-PCR primer as i5 primer (Illumina) was used for both CITE- and HASH- seq library amplification.
Following the final bead purification, all three libraries were pooled as 80% mRNA library, 10% ADT library and 10% HTO library before sequencing.
OTHER: CITE-seq (Cellular Indexing of Transcriptomes and Epitopes by sequencing)

value: Liver

tag: cell type

value: Tumor, Normal, Lymph Node and PBMC

tag: library type

value: mRNA

tag: antibodies/tags


title: scRNA-seq for WT and hm41 larval heads, 3 and 5dpf

status: Public on May 01 2023

city: Boston

first: Natasha

last: O'Brown

state: MA

address: 200 Longwood Avenue, WAB 536

department: Systems Biology

institute: Harvard Medical School

description: NovaSeq S2 kit

data_processing:
Sequencing reads were mapped to the Zebrafish GRCz11 R101 genome assembly using a custom python pipeline as previously described (see Zilionis et al., Nature Protocols 2017 and https://github.com/indrops/). Multi-seq hashtags were identified using custom code available at: https://github.com/AllonKleinLab/klunctions/tree/master/Ignas/Hashing.
We first removed background cell barcodes by only considering transcriptomes with greater than 350 UMIs for downstream analysis. In order to remove dead cells, transcriptomes were further filtered by mitochondrial read percentage (>20%). Cell demultiplexing was performed by manual inspection. Specifically, thresholds were drawn to delineate single-cells from background and multiplet populations. The resulting counts matrix was normalized to the mean UMIs per cell in the dataset.
Assembly: GRCz11
Supplementary files format and content: The gene counts matrix is an output from rsem differential gene expression analysis and is a raw counts estimate - not normalized for each gene. The first column is gene name and then after that each column represents an individual sample WT for the first 3 and then MUT for the last 3.
Supplementary files format and content: The h5ad file contains the counts matrix for the demultiplexed single-cell data. This file also holds relevant genotype and timepoint annotations as well as Multi-seq barcode counts for each cell.

organism: Danio rerio

source_name: mixed

extract_protocol:
Dissected brain tissues were dissociated using a modified protocol adapted from Bresciani et. al. 2018 (PMID: 30364607). Briefly, chemical dissociations were performed at 30.5°C using a mixture of 0.25% Trypsin-EDTA, Collagenase/Dispase (8 mg/mL) and DNaseI (20 µg/mL) for 15-20 minutes with gentle pipetting every 2.5 minutes. The dissociations were quenched using DMEM + 10% fetal bovine serum and filtered through a 40 µM cell strainer. The dissociation mixtures were spun down twice at 700g for 5 min and washed with PBS. The mixtures were then resuspended in PBS and barcoded using Multi-seq as described in McGinnis et. al. 2019 (PMID: 31209384) with slight modifications. For each sample, 80 pmoles of Lipid modified oligos (LMOs) were used to hash every 500k cells. The hashing reaction was quenched using PBS + 1% BSA. The barcoded samples were pooled into a single tube and washed twice with PBS + 1% BSA (700g for 5 min.). The pooled cell mixture was resuspended in PBS + 0.1%BSA + 18% Optiprep at a final concentration of ~300k cells/mL prior to single-cell capture with inDrops. Single-cell transcriptomes were captured by the Single-cell Core (SCC) at the Harvard Medical School as previously described (Zilionis et al., Nature Protocols 2017) using the inDrops V3 chemistry.
The Single-cell Core at the Harvard Medical School prepared the gene expression (inDrops v3 chemistry) and Multi-seq libraries.

value: mixed

tag: time

value: 3 dpf, 5 dpf


title: mRNA-derived cDNA Li124N-G


title: pooled hm41 mutant heads, 5dpf [MUT_71]

data_processing:
CLC Genomics Workbench v 11.0.1
Sequence reads were trimmed for adaptor sequence/low-quality sequence using CLC genomic benchwork (parameter- Quality limit: 0.05)
Trimmed sequence reads were mapped to GRCz11 using tophat and bowtie2 with default parameters.
Read counts were generated using rsem
Assembly: GRCz11
Supplementary files format and content: The gene counts matrix is an output from rsem differential gene expression analysis and is a raw counts estimate - not normalized for each gene. The first column is gene name and then after that each column represents an individual sample WT for the first 3 and then MUT for the last 3.
Supplementary files format and content: The h5ad file contains the counts matrix for the demultiplexed single-cell data. This file also holds relevant genotype and timepoint annotations as well as Multi-seq barcode counts for each cell.

extract_protocol:
RNA was harvested using Rneasy mini plus kit (Qiagen). 2 ug of total RNA was used for the construction of sequencing libraries.
RNA libraries for RNA-seq were prepared using Wafergen Directional RNA-Seq Library Prep Kit following manufacturer's protocols.

value: 5 dpf


title: pooled hm41 mutant heads, 5dpf [MUT_69]


title: pooled wild type heads, 5dpf [WT_67]

value: WT


title: pooled hm41 mutant heads, 5dpf [MUT_70]


title: mRNA-derived cDNA Li71P-G


title: pooled wild type heads, 5dpf [WT_66]


title: RAL195_Detrimental_HS

status: Public on Apr 25 2023

city: Salt Lake CIty

first: Katie

middle: G

last: Owings

state: UT

address: 15 North 2030 East EIHG 5200

department: Human Genetics

institute: University of Utah

data_processing:
Fastq files were trimmed by using seqtk v1.2 software
RNAseq reads were aligned to the Drosophila melanogaster reference genome (assembly BDGP6.28, Ensembl release 102) using Bowtie2 v2.2.9 software
Alignment files were sorted and converted using Samtools v1.12
Read counts were normalized using the default normalization method in DESeq2 package in R.
RAL409 was identified as an outlier and was removed for further analyses
Remaining samples were reanalyzed using Deseq2 v1.28.1
Assembly: Drosophila melanogaster reference genome (assembly BDGP6.28, Ensembl release 102)
Supplementary files format and content: resultsFDR.1_DGRP_benVdet_HS_no409.csv : Differentially expressed genes in the beneficial group (compared to detrimental group) were identified before treatment
Supplementary files format and content: resultsFDR.1_DGRP_benVdet_HS_no409.csv : Differentially expressed genes in the beneficial group (compared to detrimental group) were identified immediately post heat shock
Supplementary files format and content: counts.csv : Raw counts

organism: Drosophila melanogaster

source_name: Full body, 10 two-day old male flies

growth_protocol: Flies were maintained at 25°C on a standard diet based on the Bloomington Stock Center standard medium with malt and without soy flour. Flies were on a 12-hour light/dark cycle. For the stress preconditioning screen, DGRP strains were obtained from the Bloomington Drosophila Stock Center. Flies were 2 days old and had gone 48 hours without CO2 exposure.

extract_protocol:
Direct-zol RNA Miniprep (Zymo Research R2061) using TRIzol Reagent (ThermoFisher Cat # 15596026) and including the DNAse step
Illumina TruSeq Stranded mRNA Libraru Prep with UDI prepared and sequenced by the Huntsman Cancer Institute High-Throughput Genomics Core

treatment_protocol: The no treatment flies were frozen down at the same time as the heat shock samples. The heat shock samples from each line were heat shocked for 30 minutes at 35 ±1 °C and then frozen down immediately post heat shock.

value: Full body

value: two-day old

tag: treatment

value: Heat shock, 30 min 35+/- 1degree C


title: RAL819_Detrimental_noHS

description: resultsFDR.1_DGRP_benVdet_noHS_no409.csv


title: RAL819_Detrimental_HS


title: RAL195_Detrimental_noHS


title: pooled wild type heads, 5dpf [WT_68]


title: mRNA-derived cDNA Li15_1-G


title: RAL93_Beneficial_noHS


title: RAL69_Beneficial_noHS


title: RAL359_Beneficial_noHS


title: RAL359_Beneficial_HS


title: RAL737_Detrimental_noHS


title: RAL335_Detrimental_noHS


title: RAL409_Beneficial_HS


title: RAL737_Detrimental_HS


title: RAL387_Beneficial_HS


title: RAL387_Beneficial_noHS


title: Celf1cKO_3

status: Public on Mar 18 2023

city: Newark

first: Salil

middle: Anil

last: Lachke

state: Delaware

address: 351 Wolf Hall, University of Delaware

department: Biological Sciences

institute: University of Delaware

data_processing:
Sequence reads were trimmed to remove adaptor sequence
Trimmed reads were map to the mouse genome (GRCm38.p6) with STAR software (2.7.8a)
Uniquely mapped reads were count by featureCount (v2.0.0)
Supplementary files format and content: GRCm38.p6
Supplementary files format and content: Differential expression analysis by edgr, with cpm (count per millions) normaliezd for each sample and statistical Gene Expression difference for each genes.

organism: Mus musculus

source_name: Lens

extract_protocol:
Standard total RNA isolation protocol by RNeasy mini kit (Qiagen)
Standard protocol for Illumina sequencing

value: Lens


title: RAL304_Detrimental_noHS


title: RAL335_Detrimental_HS


title: RAL93_Beneficial_HS


title: RAL409_Beneficial_noHS


title: RAL304_Detrimental_HS


title: Celf1cKO_2


title: RAL69_Beneficial_HS


title: Celf1cKO_1


title: Control_1

description: Control


title: Control_2

description: Control


title: Control_3

description: Control


title: sample.9_1

status: Public on Feb 17 2023

city: St. Louis

first: Joseph

last: Dougherty

state: MO

address: Couch Biomedical Research Bldg 06316

department: Genetics

institute: Washington University of St. Louis

data_processing:
RNA-seq reads were aligned to the Ensembl release 101 primary assembly (GRCm38.101) with STAR version 2.7.9a.  Gene counts were derived from the number of uniquely aligned unambiguous reads by Subread:featureCount version 2.0.3. Isoform expression of known Ensembl transcripts were estimated with Salmon version 1.5.2.
All gene counts were then imported into the R/Bioconductor package EdgeR and TMM normalization size factors were calculated to adjust for samples for differences in library size.  Ribosomal genes and genes not expressed in at least ten samples greater than one count-per-million were excluded from further analysis.
The TMM size factors and the matrix of counts were then imported into the R/Bioconductor package Limma. Weighted likelihoods based on the observed mean-variance relationship of every gene and sample were then calculated for all samples and the count matrix was transformed to moderated log 2 counts-per-million with Limma’s voomWithQualityWeights.
Differential expression analysis was then performed to analyze for differences between conditions and the results were filtered for only those genes with Benjamini-Hochberg false-discovery rate adjusted p-values less than or equal to 0.05.
Assembly: Ensembl release 101 primary assembly (GRCm38.101)
Supplementary files format and content: Gene and transcript counts and CPM are provided in tab-delimited tables by their respective names. Additionally, moderated counts per million (CPM) after the filtering and quality weighting steps described are provided at the gene level, corresponding to the corrected expression values produced by limma using the linear model ~0+group, where group specified genotype_sex (thus, 10 groups altogether).

source_name: Dorsal Hippocampus

extract_protocol:
Hippocampi were lysed in 500 uls of buffer (50 mM Tris, Ph 7.4. 100 mM NaCL, 1% NP-40, supplemented with RNAsin and protease inhibitors) on ice, centrifuged at 2000g for 15 minutes at 4 ℃, and 133 µls of the supernatant was taken forward for RNA purification. This was mixed with 67 ul of Promega’s simplyRNA Tissue Kit Homogenization buffer with 1-thioglycerol (20 μL per mL), then 200 uls of Promega’s lysis, and extracted using a MaxwellRSC 48 robot (Promega) following the manufacturer’s instructions for the above Kit.
Total RNA integrity was determined using Agilent Bioanalyzer or 4200 Tapestation. Library preparation was performed with 10ng of total RNA with a Bioanalyzer RIN score greater than 8.0. ds-cDNA was prepared using the SMARTer Ultra Low RNA kit for Illumina Sequencing (Takara-Clontech) per manufacturer's protocol. cDNA was fragmented using a Covaris E220 sonicator using peak incident power 18, duty factor 20%, cycles per burst 50 for 120 seconds. cDNA was blunt ended, had an A base added to the 3' ends, and then had Illumina sequencing adapters ligated to the ends. Ligated fragments were then amplified for 12-15 cycles using primers incorporating unique dual index tags. Fragments were sequenced on an Illumina NovaSeq-6000 using paired end reads extending 150 bases.

treatment_protocol: Procedures were approved by the Institutional Animal Care and Use Committee at Washington University in St. Louis.  Adult (age range 12 - 24 weeks) wildtype and Cnih3 KO littermates, genotyped as described (Frye et al., 2021), on a C57/BL6j background were used. Mice were kept in climate-controlled facilities with a 12-hour light/dark cycle and ad libitum access to food and water.  The estrous cycle was monitored   by vaginal lavage of sterile saline for at least two consecutive days prior to tissue harvesting. Samples were allowed to dry on glass slides, rinsed with water, and stained with giemsa stain (Ricca Chemical) to improve contrast and differentiate between cell types. Vaginal cell cytological analysis was used to identify estrous cycle stage. Estrus (E) was characterized by the presence of cornified epithelial cells, metestrus (M) by a mix of leukocytes and both nucleated and cornified epithelial cells, diestrus (D) by leukocytes, and proestrus (P) by nucleated epithelial cells.  Males and females at each stage of estrous (n= 4-7 per group) were sacrificed by decapitation and dorsal hippocampi were rapidly dissected over ice, tissue was snap frozen on dry ice and stored at -80 ℃ until use.

tag: estrus

value: E

tag: age (weeks)

value: KO

value: Female


title: sample.549

value: D


title: 1076_2022_Human_Skin

status: Public on Mar 06 2023

type: RNA

city: Barcelona

first: Júlia

last: Perera-Bel

address: Doctor Aiguader, 88

department: MARData-BU

country: Spain

institute: Hospital del Mar Research Institute

hyb_protocol: cDNA was hybridized to the microarray for 16 hours at 45ºC, washing and staining of microarrays was performed using a GeneChip Fluidics Station 450 (Affymetrix, Santa Clara, CA)

scan_protocol: Arrays were scanned with GeneChip scanner GSC3000 (Affymetrix, Santa Clara, CA)

data_processing: Normalized expression signals were calculated from Affymetrix CEL files using RMA algorithm (Irizarry et al., 2003). Data was analysed with Tanscriptome Analysis Console software version 4.0.

label: Biotin

source_name: Total RNA from human skin

label_protocol: 25 ng total RNA was amplified using the TransPlex® Complete Whole Transcriptome Amplification Kit (Sigma; reference WTA2) and 8 µg of cDNA was subsequently fragmented and labeled using GeneChip Mapping 250K Nsp Assay Kit (Affymetrix; catalog # 900766), according to manufacturer's instructions.

extract_protocol: Total RNA was extracted by using TRIzol-Chloroform-Isopropanol methodology.

tag: group

value: Group 2


title: 1073_2022_Human_Skin


title: 1075_2022_Human_Skin


title: 1094_2022_Human_Skin

value: Group 1


title: sample.7_1


title: sample.643


title: sample.8_4


title: sample.6_1

value: M


title: sample.8_1


title: sample.569


title: sample.645


title: sample.8_3


title: sample.8_2

value: P


title: sample.644


title: sample.7_2


title: 1069_2022_Human_Skin


title: 1095_2022_Human_Skin


title: 1067_2022_Human_Skin


title: 1070_2022_Human_Skin


title: 1063_2022_Human_Skin


title: 1064_2022_Human_Skin


title: 1062_2022_Human_Skin


title: 1068_2022_Human_Skin


title: 1061_2022_Human_Skin


title: 1096_2022_Human_Skin


title: 1065_2022_Human_Skin


title: sample.532


title: sample.548


title: sample.5_4


title: 1066_2022_Human_Skin


title: sample.5_2


title: sample.5_3


title: sample.4_1


title: sample.5_1


title: sample.380_4


title: sample.380_1


title: sample.2589

value: male

value: Male


title: sample.2590


title: sample.3_3


title: sample.2592


title: sample.2129


title: sample.2591


title: sample.2128


title: sample.3_2


title: sample.2018


title: sample.2593


title: sample.2016


title: sample.3_1


title: sample.20_2


title: sample.2052


title: sample.2_2


title: sample.2_3


title: sample.2015


title: sample.1949


title: sample.2_1


title: sample.19_1


title: sample.14_4


title: sample.1950


title: sample.14_1


title: sample.12_5


title: sample.12_4


title: sample.12_2


title: sample.11_1


title: sample.12_3


title: sample.1_1


title: snRNA-seq of LAND-treated BJ nuclei with 10x Multiome (DEFND-seq RNA library)

status: Public on Feb 09 2023

first: Peter

middle: A

last: Sims

address: 3960 Broadway, Lasker 203AC

institute: Columbia University

data_processing:
snRNA-seq data were processed as described previously [Yuan and Sims, Scientific Reports, 2016]. The code is available at (https://github.com/simslab/DropSeqPipeline8). Briefly, we aligned the transcriptomic reads (read 2) to the human or mouse genome and transcriptome annotation using STAR v2.7.0d [Dobin et al, Bioinformatics, 2013]. For human data sets other than the mixed species experiment, we used GRCh38/Gencode v24. For the mixed species data set, we used a concatenated genome and annotation comprised of GRCh38/Gencode v32 and GRCm38/Gencode vM23. Because we are analyzing nuclei that contain a significant amount of unspliced mRNA, we identified reads that uniquely and strand-specifically aligned to the entire gene body of each gene (including exons, introns, intron-exon junctions, and exon-exon junctions), assigning an address to each aligned read containing the gene name, cell-identifying barcode sequence, and unique molecular identifier (UMI) extracted from read 1. Next, we demultiplexed the resulting table, correcting sequencing errors in the barcodes as described in Yuan and Sims [Yuan and Sims, Scientific Reports, 2016] to produce a count matrix for each sample.
snDNA-seq and snATAC-seq data were processed using a custom pipeline available here (https://github.com/simslab/dna10x). The pipeline attempts to mimic some of the basic data processing procedures that are implemented in the 10x Genomics Cell Ranger software packages for analysis of snATAC-seq and Multiome data, including the formatting of output files. We generated sample-demultiplexed fastq files using the mkfastq command in cellranger-atac v2.0.0 (10x Genomics). We aligned paired-end reads 1 and 2 to the human or mouse genome using bwa mem v0.7.17-r1188 [Li, arXiv, 2013] after removing a standard adapter sequence (CTGTCTCTTATACACATCT) from each read using cutadapt v2.8 [Martin, EMBnet.journal, 17, 1, 2011].  We then extracted all reads with an alignment score that was >90% of the read length and insert size <1 kb. We also implemented an error correction procedure for the cell-identifying barcodes from read 3. Briefly, for cell-identifying barcodes that do not appear in the standard list provided by 10x Genomics, we first determine the frequency falt of each barcode in the standard list in the data set. For each observed barcode sequence that has a Hamming distance of one from a sequence on the standard list, we estimate the posterior probability palt that the barcode deviates from the standard list due to sequencing error using the quality scores for the putatively errant base provided by Illumina or Element palt = falt10-q/10 where q is the quality score. If palt>0.975 for any alternative barcodes, then we replace the errant barcode with the alternative barcode from the standard list with the highest value of palt. Finally, we establish read addresses for each alignment that passes the filter described above comprised of a cell-identifying barcode, chromosome, a fragment start position, a fragment end position and collapse identical fragments. Finally, we sort the fragments by alignment position on the genome and output a table in the format of the “fragments.tsv” file that is typically produced by Cell Ranger.
Assembly: human-only libraries: GRCh38/Gencode v24; human/mouse mixture: GRCm38/Gencode vM23/GRCh38/Gencode v32
Supplementary files format and content: PTO016PTO021.fragments.tsv.bz2 - tab-delimited file tabulating the alignment position (chromosome, start, end), cell-identifying barcode, and read count for each unique fragment for genomic data (scATAC-seq, scDNA-seq, DEFND-seq)
Supplementary files format and content: PTO017_xSDS.fragments.tsv.bz2 - tab-delimited file tabulating the alignment position (chromosome, start, end), cell-identifying barcode, and read count for each unique fragment for genomic data (scATAC-seq, scDNA-seq, DEFND-seq)
Supplementary files format and content: PTO020.fragments.tsv.bz2 - tab-delimited file tabulating the alignment position (chromosome, start, end), cell-identifying barcode, and read count for each unique fragment for genomic data (scATAC-seq, scDNA-seq, DEFND-seq)
Supplementary files format and content: PTO018.matrix.txt.gz - tab-delimited file containing the scRNA-seq count matrix where each row is a gene (labeled by a gene ID and gene symbol) and each column is a cell (with a header containing the cell-identifying barcode and cluster number)
Supplementary files format and content: PTO025.fragments.tsv.bz2 - tab-delimited file tabulating the alignment position (chromosome, start, end), cell-identifying barcode, and read count for each unique fragment for genomic data (scATAC-seq, scDNA-seq, DEFND-seq)
Supplementary files format and content: 3T3-U87-Mix.matrix.txt.gz - tab-delimited file containing the scRNA-seq count matrix where each row is a gene (labeled by a gene ID and gene symbol) and each column is a cell (with a header containing the cell-identifying barcode and cluster number)
Supplementary files format and content: PTO034.fragments.tsv.bz2 - tab-delimited file tabulating the alignment position (chromosome, start, end), cell-identifying barcode, and read count for each unique fragment for genomic data (scATAC-seq, scDNA-seq, DEFND-seq)
Supplementary files format and content: TB5471.matrix.txt.gz - tab-delimited file containing the scRNA-seq count matrix where each row is a gene (labeled by a gene ID and gene symbol) and each column is a cell (with a header containing the cell-identifying barcode and cluster number)

source_name: BJ fibroblast cell line

growth_protocol: 3T3 cells (ATCC #CRL-1658) and U87 MG cells (ATCC #HTB-14) were cultured in Dulbecco's Modified Eagle's Medium (ATCC #30-2002) supplemented with 10% feline bovine serum (FBS, ThermoFisher # A3160401). Eagle’s Minimum Essential Medium (ATCC #30-2003) was used to culture BJ cells (ATCC CRL-2522). TrypLE Express Enzyme (ThermoFisher #12605010) was used for dissociating cells during cell passages .

extract_protocol:
snATAC-seq: Nuclei were prepared from cryopreserved BJ fibroblasts. Frozen cells were thawed and added to a Dounce homogenizer containing 1 mL of nuclei isolation buffer (NIB: 10 mM TrisHCl pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% Igepal (Millipore Sigma #18896), and 1 x protease inhibitors (Millipore Sigma #118735800001)). The sample was gently homogenized on ice with 5 strokes of the loose pestle and 5 strokes of the tight pestle. The cells were transferred to a conical and 5 mL of PBS (Gibco #10010-031) was added. The sample was spun down at 500 g for 5 minutes, washed with 10 mL of PBS and resuspended in 100 µL of 10x Genomics Nuclei Buffer (10x Genomics, #2000207).  xSDS snDNA-seq: Nuclei that were cross-linked and then treated with sodium dodecyl sulfate (SDS) to remove nucleosomes (xSDS) were prepared following Adey et al. Briefly, BJ fibroblasts were washed with 10 mL of PBS and covered in 3 mL of TrypLE (ThermoFisher #12605028) for 10 minutes at 37 °C. Then 9 mL of media was added, quenching the dissociation, and cells were spun down. The cells were resuspended in 10 mL of media with 406 µL of 37% formaldehyde (Millipore Sigma #252549) and gently rotated at room temperature for 10 minutes. 2.5 M glycine (800 µL) was then added and the suspension was incubated on ice for 5 minutes. Cells were centrifuged at 500 g for 8 minutes and washed with 10 mL of PBS. 5 mL of NIB was added to the pellet and the suspension was incubated for 20 minutes at 4 °C under rotation. Cells were then spun down at 500 g for 5 minutes and washed with 900 µL of NEBuffer 2.1 (New England Biolabs #B7202). The cells were pelleted (500 g for 5 minutes) and resuspended in NEBuffer 2.1 with 12 µL of 20% SDS. The solution was placed in a gentleMACS and vigorously shaken at 42 °C for 30 minutes. 200 µL of 10 % Triton-X (Millipore Sigma #T8787) was added and the cells were again shaken at 42 °C for 30 minutes.  LAND snDNA-seq/DEFND-seq: The LAND protocol for adherent cells using 10x reagents was adapted from the LAND protocol. BJ fibroblasts were first washed with 10 mL of PBS and dissociated with 3 mL of TrypLE for 10 minutes at 37 C. Media (9 mL) was used to quench the dissociation reaction and collect the cells. The cells were spun down (300 g for 5 minutes) and 200 µL of DEFND buffer (175 µL NIB, 10 µL 1 mg/mL protease inhibitor (Millipore Sigma #11429868001), 25 µL 100 mM lithium diiodosalicylate (Millipore Sigma #653-14-5)), was added and incubated on ice for 5 minutes. Immediately following this incubation, 10 mL of nuclei isolation buffer was added and the nuclei were centrifuged at 4 °C for 5 minutes at 500 g. The supernatant was removed and 100 µL of 10x Genomics Nuclei Buffer. A fraction of nuclei was stained with SYBR green and counted on a Countess with a GFP filter set.  GBM Cryopreserved Tissue DEFND-seq: A cryopreserved IDH wildtype Grade IV GBM sample was obtained from excess material collected for clinical purposes from de-identified brain tumor specimens. Nuclei were prepared from this anonymous sample according to XXXXX . Frozen tissue was placed in a Dounce homogenizer containing 1 mL of homogenization buffer (HB: 1 µM DTT, 250 mM sucrose, 25 mM KCl, 5 mM MgCl2, 10 mM tris, 1x protease inhibitor, 0.4 U/µL RNAse, 0.2 U/µL SUPERase In) and subjected to 15 strokes using the loose piston and 15 strokes using the tight piston. The homogenate was divided into 5 volumes and each was strained twice. The filtered homogenate was combined into a tube and centrifuged at 1000 g for 10 minutes at 4 °C. The pellet was resuspended in 1000 µL of HB and strained with 4 strainers (250 µL of homogenate for each strainer). The strained homogenate was again combined into a tube and centrifuged at 1000 g for 10 minutes at 4 °C. The pellet was resuspended in 250 µL of HB to which 250 µL of 50% iodixanol dilution media (IDM: 250 mM sucrose, 150 mM KCl, 30 mM MgCl2, 60 mM tris) was added resulting in a 25% IDM/nuclei suspension. This suspension was carefully layered on top of a tube containing 50% IDM at the bottom and 29% IDM at the surface, and centrifuged at 18,000 g for 23 minutes at 4 C. The nuclei formed a white pellet or sheet in the middle of the tube between the layers. All solution around this pellet was removed – first from the top and then from the bottom, and the pellet was resuspended in 1 mL of chilled PBS and centrifuged (500 g for 5 minutes at 4 °C). The supernatant was removed and the pellet was then treated with DEFND buffer for 5 minutes on ice. Immediately following this incubation, 5 mL of NIB with RNAse inhibitors was added at the nuclei were centrifuged at 500 g for 5 minutes at 4 °C.
snATAC-seq: Single-cell ATAC libraries were prepared with these nuclei using the 10x Genomics Single Cell ATAC kit (10x Genomics #1000176, #1000162, and #1000212) according to the manufacturer’s protocol. xSDS snDNA-seq: The cells were pelleted (500 g for 5 minutes) and resuspended in 100 µL of 10x Genomics Nuclei Buffer, counted, and tagmented following the Chromium Single Cell ATAC protocol. Tagmented nuclei were loaded onto a 10x Chromoium controller and libraries were prepared according to the Chromimum Single Cell ATAC protocol (10x Genomics #1000176, #1000162, and #1000212). LAND snDNA-seq: Nuclei were then tagmented and prepared for single-cell sequencing with the Chromium Single Cell ATAC kit (10x Genomics #1000176, #1000162, and #1000212) and protocol. DEFND-seq: Nuclei were then tagmented and prepared for single-cell sequencing with the Chromium Single Cell Multiome kit (10x Genomics #1000285, #1000230, #1000215).

tag: cell line

value: BJ fibroblast cell line

value: fibroblasts


title: snDNA-seq of LAND-treated BJ nuclei with 10x Multiome (DEFND-seq DNA library)

library_source: genomic

molecule: genomic DNA


title: snDNA-seq of xSDS-treated BJ nuclei with 10x scATAC-seq kit


title: snATAC-seq of BJ nuclei with 10x scATAC-seq


title: snRNA-seq of LAND-treated glioblastoma nuclei with 10x Multiome (DEFND-seq RNA library)

source_name: glioblastoma tissue

value: glioblastoma tissue


title: STR3_Sotalol_Control_vHC_Rep_8

status: Public on Feb 02 2023

city: Zürich

first: Lukas

middle: Matthias

last: von Ziegler

address: Winterthurerstrasse 190

department: D-HESZ

country: Switzerland

institute: ETH Zürich

data_processing:
Pseudoalignment using kallisto (version 0.44.0) on the GENCODE M17 transcriptome
Aggregation on the protein coding level
Assembly: GRCm38
Supplementary files format and content: .tar.gz zipped directories with kallisto output (pseudoaligned read counts and tpm measures of trancripts) containing .h5 and .tsv files

source_name: brain

extract_protocol:
Trizol
illumina TrueSeq

value: brain

tag: brainregion

value: hippocampus

tag: subregion

value: vHC

tag: animalid

tag: replicate

tag: strain

value: Control

tag: injection

value: Sotalol


title: snRNA-seq of LAND-treated U87 and 3T3 nuclei with 10x Multiome (DEFND-seq RNA library)

organism:
Homo sapiens
Mus musculus

source_name: U87 human glioma cell line / 3T3 mouse fibroblast cell line

value: human glioma cells / mouse fibroblasts


title: sample.10_1

value: KO

tag: Sex

value: Female


title: STR3_Propranolol_Control_vHC_Rep_8

tag: tissue

value: Propranolol

value: female


title: STR3_Saline_Stress_vHC_Rep_8

value: Stress

value: Saline


title: STR3_Sotalol_Stress_vHC_Rep_8

value: male


title: STR3_Saline_Control_vHC_Rep_8


title: snDNA-seq of LAND-treated U87 and 3T3 nuclei with 10x Multiome (DEFND-seq DNA library)

value: U87 human glioma cell line / 3T3 mouse fibroblast cell line

tag: cell type


title: STR3_Saline_Stress_vHC_Rep_7


title: sample.14_2

value: P

tag: genotype


title: STR3_Propranolol_Stress_vHC_Rep_7


title: STR3_Saline_Control_vHC_Rep_7


title: STR3_Propranolol_Control_vHC_Rep_7


title: sample.13_1

value: M

value: WT


title: STR3_Sotalol_Control_vHC_Rep_7

value: brain


title: sample.12_1

value: E


title: STR3_Propranolol_Control_vHC_Rep_6

value: hippocampus

value: vHC


title: snDNA-seq of LAND-treated BJ nuclei with 10x scATAC-seq kit

value: BJ fibroblast cell line

value: fibroblasts


title: STR3_Sotalol_Stress_vHC_Rep_6

value: Sotalol


title: STR3_Propranolol_Stress_vHC_Rep_6


title: STR3_Saline_Control_vHC_Rep_6

value: Control


title: STR3_Saline_Stress_vHC_Rep_6


title: STR3_Propranolol_Stress_vHC_Rep_8

value: Propranolol


title: STR3_Saline_Stress_vHC_Rep_5

value: female


title: STR3_Sotalol_Control_vHC_Rep_6


title: STR3_Sotalol_Control_vHC_Rep_5


title: STR3_Saline_Control_vHC_Rep_4


title: STR3_Saline_Stress_vHC_Rep_4

value: Stress


title: STR3_Saline_Control_vHC_Rep_5

value: Saline


title: STR3_Propranolol_Stress_vHC_Rep_5


title: STR3_Sotalol_Stress_vHC_Rep_5

value: male


title: STR3_Saline_Control_vHC_Rep_3


title: STR3_Propranolol_Stress_vHC_Rep_4


title: STR3_Propranolol_Control_vHC_Rep_4


title: STR3_Sotalol_Stress_vHC_Rep_7


title: STR3_Sotalol_Stress_vHC_Rep_3


title: STR3_Saline_Stress_vHC_Rep_3


title: STR3_Sotalol_Control_vHC_Rep_3


title: STR3_Propranolol_Stress_vHC_Rep_3


title: STR3_Propranolol_Control_vHC_Rep_3


title: STR3_Sotalol_Control_vHC_Rep_4


title: STR3_Propranolol_Control_vHC_Rep_2


title: STR3_Propranolol_Stress_vHC_Rep_2


title: STR3_Propranolol_Stress_vHC_Rep_1


title: STR3_Saline_Control_vHC_Rep_2


title: STR3_Sotalol_Stress_vHC_Rep_2


title: STR3_Saline_Stress_vHC_Rep_2


title: STR3_Sotalol_Control_vHC_Rep_2


title: STR3_Propranolol_Control_vHC_Rep_5


title: STR3_Saline_Control_vHC_Rep_1


title: STR3_Saline_Stress_vHC_Rep_1


title: STR3_Propranolol_Control_vHC_Rep_1


title: STR2_Propanolol_Stress_vHC_Rep_5

value: Propanolol


title: STR3_Sotalol_Stress_vHC_Rep_4


title: STR2_Saline_Control_vHC_Rep_5


title: STR2_Saline_Stress_vHC_Rep_6


title: STR2_Saline_Yohimbine_vHC_Rep_5

value: Yohimbine


title: STR2_Prazosin_Stress_vHC_Rep_5

value: Prazosin


title: STR2_Propanolol_Stress_vHC_Rep_4


title: STR2_Prazosin_Stress_vHC_Rep_6


title: STR2_Saline_Yohimbine_vHC_Rep_4


title: STR2_Prazosin_Stress_vHC_Rep_4


title: STR2_Saline_Control_vHC_Rep_4


title: STR2_Saline_Stress_vHC_Rep_4


title: STR2_Propanolol_Stress_vHC_Rep_3


title: STR2_Saline_Yohimbine_vHC_Rep_3


title: STR2_Propanolol_Stress_vHC_Rep_2


title: STR2_Saline_Stress_vHC_Rep_3


title: STR2_Saline_Control_vHC_Rep_3


title: STR3_Sotalol_Control_vHC_Rep_1


title: STR2_Saline_Yohimbine_vHC_Rep_2


title: STR2_Prazosin_Stress_vHC_Rep_2


title: STR2_Propanolol_Stress_vHC_Rep_1


title: STR3_Sotalol_Stress_vHC_Rep_1


title: STR2_Saline_Stress_vHC_Rep_2


title: STR2_Saline_Control_vHC_Rep_2


title: STR2_Saline_Stress_vHC_Rep_5


title: STR2_Saline_Yohimbine_vHC_Rep_1


title: STR2_Prazosin_Stress_vHC_Rep_1


title: STR2_Saline_Yohimbine_vHC_Rep_6


title: STR2_Propanolol_Stress_dHC_Rep_6

value: dHC


title: STR2_Saline_Stress_dHC_Rep_6

value: brain


title: STR2_Saline_Control_vHC_Rep_1


title: STR2_Saline_Yohimbine_dHC_Rep_6


title: STR2_Saline_Stress_vHC_Rep_1


title: STR2_Prazosin_Stress_vHC_Rep_3


title: STR2_Prazosin_Stress_dHC_Rep_5


title: STR2_Propanolol_Stress_dHC_Rep_5


title: STR2_Saline_Control_dHC_Rep_5


title: STR2_Saline_Yohimbine_dHC_Rep_4


title: STR2_Saline_Stress_dHC_Rep_5


title: STR2_Saline_Stress_dHC_Rep_4


title: STR2_Prazosin_Stress_dHC_Rep_4


title: STR2_Propanolol_Stress_dHC_Rep_4


title: STR2_Saline_Stress_dHC_Rep_3


title: STR2_Propanolol_Stress_dHC_Rep_3


title: STR2_Saline_Control_dHC_Rep_4


title: STR2_Saline_Yohimbine_dHC_Rep_3


title: STR2_Propanolol_Stress_dHC_Rep_2


title: STR2_Saline_Control_dHC_Rep_6


title: STR2_Prazosin_Stress_dHC_Rep_3


title: STR2_Saline_Yohimbine_dHC_Rep_2


title: STR2_Saline_Yohimbine_dHC_Rep_1


title: STR2_Saline_Control_dHC_Rep_2


title: STR2_Saline_Control_dHC_Rep_3


title: STR2_Saline_Stress_dHC_Rep_2


title: STR2_Prazosin_Stress_dHC_Rep_2


title: STR2_Saline_Yohimbine_dHC_Rep_5


title: STR2_Prazosin_Stress_dHC_Rep_6


title: STR2_Saline_Stress_dHC_Rep_1


title: STR2_Prazosin_Stress_dHC_Rep_1


title: STR2_Saline_Control_dHC_Rep_1


title: STR2_Propanolol_Stress_dHC_Rep_1


title: 112

status: Public on Jan 17 2023

hyb_protocol: Sample was hybridized to Affymetrix miRNA 4.0 array using an Affymetrix GeneChip Hybridization Oven 645

scan_protocol: GeneChip was scanned using the Affymetrix Expression Wash, Stain and Scan User Manual (P/N 702731 Rev. 3)  (Affymetrix Inc., Santa Clara, CA, USA) using Affymetrix GeneChip Scanner 3000 7G.

data_processing: The data were analyzed using the RMA algorithm and then LIMMA was applied to calculate significant differential expression between samples

source_name: microRNA from epithelial components of cutaneous squamous cell carcinoma (SCC)

label_protocol: 750ng of total RNA from each sample was processed and labelled according to manual FlashTag™ Biotin HSR RNA Labeling Kit (P/N 703095, Rev.2) (Affymetrix Inc., Santa Clara, CA, USA)

extract_protocol: microRNA was extracted following Qiagen protocol

tag: tissue/condition

value: Ctrl; epithelial components of cutaneous squamous cell carcinoma (SCC)

tag: batch

tag: molecule type

value: microRNA


title: 129


title: 132


title: 101


title: 108


title: 121


title: 92


title: 123


title: 128


title: 99


title: 68

source_name: microRNA from epithelial components of cutaneous squamous cell carcinoma (SCC)  in metastatic progression


title: 64

value: epithelial components of cutaneous squamous cell carcinoma (SCC)  in metastatic progression


title: 42


title: 87


title: 36


title: 91


title: 29


title: 32


title: 16


title: 12


title: 12ADKCi_1

description: Gene expression data from human SCC cell line (DKCi)

hyb_protocol: Sample was hybridized to Affymetrix Clariom S Human array using an Affymetrix GeneChip Hybridization Oven 645

source_name: human SCC cell line (DKCi)

label_protocol: 100ng of total RNA from each sample was processed and labelled according to manual GeneChip WT PLUS Reagent kit (P/N 703174 Rev. 2, Affymetrix Inc., Santa Clara, CA, USA)

extract_protocol: Total RNA was extracted following Qiagen protocol


title: 26


title: A2780 TK+ rep 1

status: Public on Jan 09 2023

city: New Haven

first: Jason

last: Sheltzer

state: CT

address: 300 George Street

department: Surgery

institute: Yale University

description: Ovarian cancer cell line, harboring gain of chr1q and TK+ integrant

data_processing:
Custom Novogene QC scripts
Novogene: Remove reads containing adapters
Novogene: Remove reads containing >10% undetermined bases (N)
Novogene: Remove reads with >=50% of bases with Qscore <= 5
Hisat2 (v2.0.5) alignment to hg38
featureCounts v1.5.0-p3 to count reads mapped to each gene
Assembly: hg38
Supplementary files format and content: tab-delimited text files include FPKM values for each Sample

growth_protocol: Cells were plated and allowed to grow for 24 hours prior to extraction

extract_protocol:
Qiagen RNEeasy
In brief, mRNA was purified from total cellular RNA using poly-T oligo-attached magnetic beads. After fragmentation, first strand cDNA was synthesized using random hexamer primers, followed by second strand cDNA synthesis. Subsequently, libraries were prepared with end repair, A-tailing, adapter ligation, size selection, amplification, and purification.

treatment_protocol: Single cell derived clones with engineered chr losses were generated through a collection of ReDACT techniques

value: ovarian cancer

value: TK+

tag: treatment

value: TK+ integrant


title: 12ADKCi_2


title: 1A


title: A2780 WT rep 2

description: Ovarian cancer cell line, harboring gain of chr1q

value: WT

value: Wildtype


title: 12AGi_1

description: Gene expression data from human SCC cell line (CTRL)

source_name: human SCC cell line (CTRL)

value: CTRL


title: 12AGi_2


title: 12AGi_3


title: A2780 1q-loss c6 rep 1

description: Ovarian cancer cell line with engineered loss of chr1q

value: ReDACT-NS, targeting chr1q


title: A2780 1q-loss c6 rep 2


title: A2780 1q-loss c5 rep 2


title: A2780 TK+ rep 2


title: A2780 1q-loss c2 rep 1


title: A2780 1q-loss c5 rep 1


title: 12ADKCi_3


title: EF_Sarcoma_2 [10_30245_GATCAG]

status: Public on Dec 31 2022

data_processing:
Alignment was made using STAR version 2.6.1b
The table of counts was obtained with FeatureCounts function in the package subread, version 1.5
Genome_build: hg38
Supplementary_files_format_and_content: counts

source_name: Sarcoma derived from hMSLCs  expressing EWSFLI1

growth_protocol: Human Mesenchymal Stem Like Cells injected into the gastrocnemius muscles of NOD/SCID mice

extract_protocol:
RNA was extracted following Sigma-Aldrich protocol
TruSeq stranded mRNA (Illumina)

tag: host strain

value: NOD/SCID mice

tag: cell type source

value: human Mesenchymal Stem Like Cells  (hMSLCs) expressing EWSFLI1

value: Sarcoma

value: EWSRndFLI


title: EF_Lymphoma_1 [1_30239_ACAGTG]

source_name: Lymphoma derived from hMSLCs expressing EWSFLI1

value: Lymphoma


title: Control Lymphoma_2 [5_30241_CTTGTA]

source_name: Lymphoma derived from control hMSLCs

value: control human Mesenchymal Stem Like Cells (hMSLCs)


title: Control Lymphoma_1 [3_30240_GCCAAT]


title: EF_Sarcoma_1 [8_30243_AGTTCC]


title: hNSC_H2O2cGP2

city: Tsukuba

first: Kazutoshi

last: Murotomi

address: 1-1-1 Higashi

institute: Advanced Industrial Science and Technology

data_processing:
Sequencing data were analyzed using an open bioinformatics tool for the quick analysis of RNA-seq data RaNA-seq
FASTQ files are pre-processed with the Fastp tool, and the expression quantification is performed with Salmon
The quality of the raw date was represented some graphs produced by the package RJSplot
the distances between plots in the graphs are calculated with SERE
DEseq2 was used to identify the differentially expressed genes (DEGs) with more than two-fold changes in transcripts per million (TPM) between control and cGP, H2O2, or H2O2+cGP-treated groups
Genome_build: hg19
Supplementary_files_format_and_content: Excel files include differential expression analysis for comparision between two exerimental groups. Expression levels in mRNA are calculated as TPMs values using salmon (Patro et al., Nat Methods., 2017)
Supplementary_files_format_and_content: DE_cont_vs_cGP.xlsx: Comparision of abundance measurements between control and cGP
Supplementary_files_format_and_content: DE_cont_vs_H2O2.xlsx: Comparision of abundance measurements between control and H2O2
Supplementary_files_format_and_content: DE_cont_vs_H2O2+cGP.xlsx: Comparision of abundance measurements between control and H2O2+cGP
Supplementary_files_format_and_content: DE_H2O2_vs_H2O2+cGP.xlsx: Comparision of abundance measurements between H2O2 and H2O2+cGP

source_name: Neural stem cells

extract_protocol:
Trizol reagents were used for the extraction of total RNA.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: cGP and/or H2O2 treated to hfNSCs for 4h

value: Human fetal neural stem cells isolated from human cerebral cortex from male fetus at embryonic week 14

value: 100uM H2O2 + 1 mM cGP for 4 h


title: hNSC_H2O2_3

value: 100uM H2O2 for 4 h


title: A2780 WT rep 1

tag: cell line


title: hNSC_H2O2cGP3


title: hNSC_H2O2_1


title: hNSC_H2O2cGP1


title: A2780 1q-loss c2 rep 2


title: hNSC_cont2

value: Non-treatment


title: hNSC_cGP3

value: 1 mM cGP for 4 h


title: EF_Lymphoma_2 [6_30242_GTGAAA]


title: hNSC_cGP2


title: hNSC_cont3


title: EF_hMSLC1_1

hyb_protocol: Sample was hybridized to Affymetrix Human Gene 2.0 ST array using an Affymetrix GeneChip Hybridization Oven 645

scan_protocol: GeneChip was scanned using the Affymetrix Expression Wash, Stain and Scan User Manual (P/N 702731 Rev. 3. Affymetrix Inc., Santa Clara, CA, USA) using Affymetrix GeneChip Scanner 3000 7G

label: biotin

source_name: human Mesenchymal Stem Like Cell Line 1 expressing EWSFLI1

extract_protocol: RNA was extracted following Sigma-Aldrich protocol

value: Mesenchymal Stem Like Cells

tag: genotype/variation

value: ectopic EWSR1/FLI1 expression


title: EF_hMSLC2_1

source_name: human Mesenchymal Stem Like Cell Line 2 expressing EWSFLI1


title: hNSC_cont1


title: CONTROL_hMSLC_2

source_name: Control human Mesenchymal Stem Like Cells

value: hMSLC

value: CONTROL


title: CONTROL_hESC_2

source_name: Control human Embryonic Stem Cells

value: hESC

value: Embryonic Stem Cells


title: CONTROL_hMSLC_1


title: EF_hMSLC1_2


title: hNSC_cGP1


title: CONTROL_hESC_1


title: EF_hMSLC2_2


title: EF_hESC_2

source_name: human Embryonic Stem Cells expressing EWSFLI1


title: LPS group lung tissue [LPS-6]

status: Public on Dec 01 2022

city: Kaifeng

first: Huanling

last: Yao

state: Henan

address: Jin Ming Avenue

department: School of Life Sciences

country: China

institute: Henan University

data_processing:
Image analysis and base calling were conducted by the NovaSeq Control Software (NCS) + OLB + GAPipeline-1.6 (Illumina) on the NovaSeq instrument.
Sequence reads were filtered for quality using the fastqc v 0.11.9 and the trim_galore softwares v 0.6.7 (parameter---length 36,--max_n 3)
Trimmed sequence reads were mapped to the ENSEMBL reference genome Mus_musculus GRCm39 using the Hisat2 software v2.2.1
Gene expression quantity were analyzed by the featureCounts software v2.0.1.
Assembly: the ENSEMBL reference genome Mus_musculus GRCm39
Supplementary files format and content: tab-delimited text files include raw counts for each Sample

source_name: lung tissue

extract_protocol:
Total RNA of each sample was extracted using TRIzol Reagent/RNeasy Mini Kit (Qiagen)/other kits.
Next generation sequencing library preparations were constructed according to the manufacturer’s protocol. The poly(A) mRNA isolation was performed using Poly(A) mRNA Magnetic Isolation Module or rRNA removal Kit. The mRNA fragmentation and priming was performed using First Strand Synthesis Reaction Buffer and Random Primers. First strand cDNA was synthesized using ProtoScript II Reverse Transcriptase and the second-strand cDNA was synthesized using Second Strand Synthesis Enzyme Mix. The purified double-stranded cDNA by beads was then treated with End Prep Enzyme Mix to repair both ends and add a dA-tailing in one reaction, followed by a T-A ligation to add adaptors to both ends. Size selection of Adaptor-ligated DNA was then performed using beads, and fragments of ~400 bp (with the approximate insert size of 300 bp) were recovered. Each sample was then amplified by PCR using P5 and P7 primers, with both primers carrying sequences which can anneal with flow cell to perform bridge PCR andP5/ P7 primer carrying index allowing for multiplexing. The PCR products were cleaned up using beads, validated using an Qsep100 (Bioptic, Taiwan, China), and quantified by Qubit3.0 Fluorometer (Invitrogen, Carlsbad, CA, USA).

value: lung tissue

value: LPS


title: cerebellar endothelial cells wild-type 3

status: Public on Nov 22 2022

city: Uppsala

first: Ying

last: Sun

address: Dag Hammarskjölds väg 20

country: Sweden

institute: Uppsala University

description:
Sample 3
cerebellar endothelial cells from wild-type mouse 3 at postnatal day 9

data_processing:
The quality control and prepossessing of the raw reads were done using Trim Galore.
Reads were mapped to the GRCm38 mouse genome using default settings of STAR (version 2.5.3a).
The number of reads per gene were counted while mapping by STAR.
Reads normalization was done using the DESeq2 Bioconductor package.
Assembly: GRCm38
Supplementary files format and content: Excel file with normalized counts for each sample

source_name: cerebellar endothelial cells

extract_protocol:
Endothelial cells were isolated from the cerebella of 3 wild-type and 3 Ccm3 knock-out mice with the Adult Brain Dissociation kit (130-107-677; MiltenyiBiotech), CD45 MicroBeads (30-052-301; MiltenyiBiotech) and CD31 MicroBeads (30-097-418; MiltenyiBiotech) according to the manufacturer’s instructions. Total RNA was extracted from each sample with the RNeasy Micro Kit (Qiagen) in the automated QIACube (Qiagen) platform.
RNA sequencing libraries were prepared with SMARTer Stranded Total RNA Sample Prep kit - Mammalian kit (Clontech), according to the manufacturer’s instructions. This kit includes depletion of ribosomal RNA prior to library synthesis, barcoding and amplification. 1 μl of each library was analyzed in the Agilent 2100 bioanalyzer for validation of fragment size and quality control of the amplified RNA-seq library, using the High Sensitivity DNA kit (Agilent).

value: wild-type

value: brain

value: cerebellar endothelial cells

tag: age

value: postnatal day 9


title: cerebellar endothelial cells Ccm3 knock-out 1

description:
Sample 4
cerebellar endothelial cells from Ccm3 knock-out mouse 1 at postnatal day 9

value: Ccm3 knock-out


title: cerebellar endothelial cells wild-type 2

description:
Sample 2
cerebellar endothelial cells from wild-type mouse 2 at postnatal day 9


title: early gastrula stage  [PmEG]

status: Public on Nov 16 2022

city: Providence

first: Gary

last: Wessel

state: RI

address: 185 Meeting St

department: Molecular Biology, Cell Biology & Biochemistry

institute: Brown University

data_processing:
Single cell unique molecular identifier (UMI) counting (counting of unique barcodes given to individual transcript molecules), was performed using Cell Ranger Single Cell Software Suite 3.0.2 from 10X Genomics. The custom transcriptome reference was generated from P. miniata assembly V2.0 (echinobase.org) using CellRanger mkref. The annotation for PmNanos was manually edited to account for a longer 3’UTR. Duplicate blastula and gastrula stage libraries were aggregated using the cellranger aggr function. Cellranger gene expression matrices were further analyzed using the R package Seurat v 3.1.4 (Butler et al., 2019). Cells of 8hpf to 14hpf stages with at least 1000 and at most 6000 genes (features), and cells with at least 1000 and at most 2500 genes in blastula to mid-gastrula stages were included in downstream analysis. Individual datasets were normalized by scaling gene expression in each cell by total gene expression and then log transformed. The top 2000 highly variable genes across the datasets were then used to integrate the datasets. Individual time point datasets were integrated using the Seurat toolkit Harmony to identify conserved cell populations across the datasets. The clustering parameters used were dimensions: 20, resolution: 1.0. Cluster markers were found using FindConservedMarkers and FindMarkers functions.
Aggregate technical replicates of  blastula (B), early gastrula (EG), and  mid-gastrula (MG) using cellanger aggr.
Genome_build: P. miniata assembly V2.0 (echinobase.org)
Supplementary_files_format_and_content: tab-delimited text files include matrices that contain UMI-corrected counts for each cell and integrated seurat object

organism: Patiria miniata

source_name: embryo

extract_protocol: Single cell encapsulation was performed using the Chromium Single Cell Chip B kit on the 10x Genomics Chromium Controller. Single cell cDNA and libraries were prepared using the Chromium Single Cell 3’ Reagent kit v3 Chemistry. Libraries were sequenced by Genewiz on the Illumina Hiseq (2x150 bp paired-end runs).

tag: Stage

value: early gastrula stage

value: embryo


title: cerebellar endothelial cells Ccm3 knock-out 2

description:
Sample 5
cerebellar endothelial cells from Ccm3 knock-out mouse 2 at postnatal day 9


title: hNSC_H2O2_2

value: Human fetal neural stem cells isolated from human cerebral cortex from male fetus at embryonic week 14

value: 100uM H2O2 for 4 h


title: cerebellar endothelial cells Ccm3 knock-out 3

description:
Sample 6
cerebellar endothelial cells from Ccm3 knock-out mouse 3 at postnatal day 9


title: blastula stage [PmB]

value: blastula stage


title: 10hpf [Pm10h]


title: cerebellar endothelial cells wild-type 1

description:
Sample 1
cerebellar endothelial cells from wild-type mouse 1 at postnatal day 9


title: mid-gastrula stage [PmMG]

value: mid-gastrula stage


title: EF_hESC_1

value: ectopic EWSR1/FLI1 expression


title: 14 hpf [Pm14h]

value: 14 hpf


title: 8 hpf [Pm8h]

value: 8 hpf


title: SH276

status: Public on Oct 17 2022

description: Gene expression data

scan_protocol: GeneChip was scanned using the Affymetrix Expression Wash, Stain and Scan User Manual (P/N 702731 Rev. 3)  (Affymetrix Inc., Santa Clara, CA, USA) using Affymetrix GeneChip Scanner 3000 7G

source_name: Total RNA from peripheral blood

label_protocol: a total of 100ng from each sample was processed and labelled according to manual GeneChip WT PLUS Reagent kit (P/N 703174 Rev. 2, Affymetrix Inc., Santa Clara, CA, USA)

extract_protocol: Total RNA was isolated from whole blood samples using the PaxGene™ blood RNA system (PreAnalytiX, Qiagen/Becton Dickson) according to the manufacturer specifications

value: peripheral blood

tag: patient diagnosis

value: community-acquired pneumonia (CAP)

tag: hospital

tag: mortality


title: RP340

description: Gene expression data

tag: Sex


title: RF267

description: Gene expression data

tag: tissue


title: SR312

description: Gene expression data


title: RT314

description: Gene expression data


title: RV249

description: Gene expression data


title: ST195

description: Gene expression data


title: SA238

description: Gene expression data


title: RC289

description: Gene expression data


title: RP261

description: Gene expression data


title: VG328

description: Gene expression data


title: VC189

description: Gene expression data


title: SG296

description: Gene expression data


title: TG198

description: Gene expression data


title: PM302

description: Gene expression data


title: RA245

description: Gene expression data


title: PL190

description: Gene expression data


title: PP310

description: Gene expression data


title: RR283

description: Gene expression data


title: PG287

description: Gene expression data


title: MP229

description: Gene expression data


title: MR281

description: Gene expression data


title: MR272

description: Gene expression data


title: PG341

description: Gene expression data


title: RS339

description: Gene expression data


title: MO298

description: Gene expression data


title: MI197

description: Gene expression data


title: NG293

description: Gene expression data


title: MO278

description: Gene expression data


title: MO199

description: Gene expression data


title: RL271

description: Gene expression data


title: MP200

description: Gene expression data


title: MM233

description: Gene expression data


title: MM303

description: Gene expression data


title: MC327

description: Gene expression data


title: MM232

description: Gene expression data


title: MJ333

description: Gene expression data


title: ML313

description: Gene expression data


title: MN208

description: Gene expression data


title: ME210

description: Gene expression data


title: MM315

description: Gene expression data


title: MA337

description: Gene expression data


title: ME349

description: Gene expression data


title: MM225

description: Gene expression data


title: MA305

description: Gene expression data


title: MG320

description: Gene expression data


title: LR336

description: Gene expression data


title: MC224

description: Gene expression data


title: LP212

description: Gene expression data


title: MB275

description: Gene expression data


title: MA251

description: Gene expression data


title: MA237

description: Gene expression data


title: LE291

description: Gene expression data


title: JT280

description: Gene expression data


title: JV292

description: Gene expression data


title: JR250

description: Gene expression data


title: JS274

description: Gene expression data


title: LB193

description: Gene expression data


title: JR347

description: Gene expression data


title: JP255

description: Gene expression data


title: JO300

description: Gene expression data


title: JM262

description: Gene expression data


title: JM231

description: Gene expression data


title: JM221

description: Gene expression data


title: JL307

description: Gene expression data


title: JL290

description: Gene expression data


title: JD335

description: Gene expression data


title: JG324

description: Gene expression data


title: JL277

description: Gene expression data


title: JS239

description: Gene expression data


title: JD343

description: Gene expression data


title: JM214

description: Gene expression data


title: JM273

description: Gene expression data


title: JA234

description: Gene expression data


title: JC260

description: Gene expression data


title: JC252

description: Gene expression data


title: JC294

description: Gene expression data


title: JB257

description: Gene expression data


title: JB306

description: Gene expression data


title: IS226

description: Gene expression data


title: JB282

description: Gene expression data


title: HF244

description: Gene expression data


title: HB195

description: Gene expression data


title: IG311

description: Gene expression data


title: HB167

description: Gene expression data


title: HB212

description: Gene expression data


title: HB146

description: Gene expression data


title: HC323

description: Gene expression data


title: HB181

description: Gene expression data


title: HB165

description: Gene expression data


title: IO240

description: Gene expression data


title: HB141

description: Gene expression data


title: HB169

description: Gene expression data


title: HB163

description: Gene expression data


title: HB140

description: Gene expression data


title: HB144

description: Gene expression data


title: HB159

description: Gene expression data


title: HB215

description: Gene expression data


title: HB136

description: Gene expression data


title: HB142

description: Gene expression data


title: HB137

description: Gene expression data


title: HB143

description: Gene expression data


title: HB132

description: Gene expression data


title: HB138

description: Gene expression data


title: HB122

description: Gene expression data


title: HB123

description: Gene expression data


title: HB133

description: Gene expression data


title: HB116

description: Gene expression data


title: HB113

description: Gene expression data


title: HB102

description: Gene expression data


title: FV213

description: Gene expression data


title: HB121

description: Gene expression data


title: HB101

description: Gene expression data


title: FV247

description: Gene expression data


title: GB338

description: Gene expression data


title: FO329

description: Gene expression data


title: HB180

description: Gene expression data


title: FP301

description: Gene expression data


title: FR204

description: Gene expression data


title: FG348

description: Gene expression data


title: HB112

description: Gene expression data


title: FL228

description: Gene expression data


title: FM317

description: Gene expression data


title: HB99

description: Gene expression data


title: FL192

description: Gene expression data


title: FB216

description: Gene expression data


title: FG191

description: Gene expression data


title: EP344

description: Gene expression data


title: EP331

description: Gene expression data


title: EG254

description: Gene expression data


title: HB98

description: Gene expression data


title: HB135

description: Gene expression data


title: EV201

description: Gene expression data


title: DL202

description: Gene expression data


title: EA241

description: Gene expression data


title: DT322

description: Gene expression data


title: EA230

description: Gene expression data


title: EC345

description: Gene expression data


title: CM256

description: Gene expression data


title: DC265

description: Gene expression data


title: CG248

description: Gene expression data


title: CB263

description: Gene expression data


title: CC284

description: Gene expression data


title: CA285

description: Gene expression data


title: CA269

description: Gene expression data


title: CC304

description: Gene expression data


title: B222

description: Gene expression data


title: DB342

description: Gene expression data


title: CA227

description: Gene expression data


title: B210

description: Gene expression data


title: B219

description: Gene expression data


title: B189

description: Gene expression data


title: B204

description: Gene expression data


title: B190

description: Gene expression data


title: B155

description: Gene expression data


title: B187

description: Gene expression data


title: B178

description: Gene expression data


title: B171

description: Gene expression data


title: B179

description: Gene expression data


title: B176

description: Gene expression data


title: B175

description: Gene expression data


title: B152

description: Gene expression data


title: B202

description: Gene expression data


title: B153

description: Gene expression data


title: B151

description: Gene expression data


title: B124

description: Gene expression data


title: B150

description: Gene expression data


title: B149

description: Gene expression data


title: B108

description: Gene expression data


title: B128

description: Gene expression data


title: B97

description: Gene expression data


title: B134

description: Gene expression data


title: B130

description: Gene expression data


title: B129

description: Gene expression data


title: B148

description: Gene expression data


title: AZ330

description: Gene expression data


title: B126

description: Gene expression data


title: B104

description: Gene expression data


title: B107

description: Gene expression data


title: AR196

description: Gene expression data


title: AV334

description: Gene expression data


title: HB117

description: Gene expression data


title: AV318

description: Gene expression data


title: AP279

description: Gene expression data


title: AP266

description: Gene expression data


title: AM205

description: Gene expression data


title: LLC1 - cisplatine+KW_rep5

status: Public on Sep 09 2022

city: Lille

first: David

last: Blum

state: Cedex

address: 1 place de Verdun

department: Lille Neurosience & Cognition

country: France

institute: Inserm

description: c761_LLC1 - cisplatine+KW_rep5 (raw)

data_processing:
Image analysis and base calling were performed using RTA 2.7.3 and bcl2fastq 2.17.1.14.
Reads were mapped onto the mm10 assembly of Mus musculus genome using STAR [3] version 2.5.3a.
Gene expression quantication was performed from uniquely aligned reads using htseq-count version  0.6.1p1, with annotations from Ensembl version 90 and union mode.
Genome_build: mm10
Supplementary_files_format_and_content: Tab-separated text file with raw read counts per Ensembl Gene (ensembl v102).

source_name: tumors

extract_protocol:
Total RNA was extracted by phenol/ chloroform and subsequently precipitated in isopropanol as described previously (Vomelováet al., 2009). Total RNAs from cultured renal cells were extracted usingRNeasy Mini kit (Qiagen) following manufacturer’s recommendations.
RNA-Seq libraries were generated from 350 ng of total RNA using TruSeq Stranded mRNA Library Prep Kit and TruSeq RNA Single Indexes kits A and B (Illumina, San Diego, CA), according to manufacturer's instructions. Briefly, following purification with poly-T oligo attached magnetic beads, the mRNA was fragmented using divalent cations at 94oC for 2 minutes. The cleaved RNA fragments were copied into first strand cDNA using reverse transcriptase and random primers. Strand specificity was achieved by replacing dTTP with dUTP during second strand cDNA synthesis using DNA Polymerase I and RNase H. Following addition of a single 'A' base and subsequent ligation of the adapter on double stranded cDNA fragments, the products were purified and enriched with PCR (30 sec at 98oC; [10 sec at 98oC, 30 sec at 60oC, 30 sec at 72oC] x 12 cycles; 5 min at 72oC) to create the cDNA library. Surplus PCR primers were further removed by purification using AMPure XP beads (Beckman-Coulter, Villepinte, France) and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis.

value: LLC1 tumor

tag: agent

value: cisplatine+KW


title: LLC1 - cisplatine+KW_rep1

description: c734_LLC1 - cisplatine+KW_rep1 (raw)


title: LLC1 - cisplatine+KW_rep3

description: c736_LLC1 - cisplatine+KW_rep3 (raw)


title: AP308

description: Gene expression data


title: AM219

description: Gene expression data


title: LLC1 - cisplatine_rep4

description: c758_LLC1 - cisplatine_rep4 (raw)

value: cisplatine


title: LLC1 - cisplatine_rep3

description: c757_LLC1 - cisplatine_rep3 (raw)


title: LLC1 - cisplatine+KW_rep2

description: c735_LLC1 - cisplatine+KW_rep2 (raw)


title: LLC1 - KW_rep2

description: c730_LLC1 - KW_rep2 (raw)

value: KW


title: LLC1 - KW_rep4

description: c754_LLC1 - KW_rep4 (raw)


title: LLC1 - KW_rep5

description: c756_LLC1 - KW_rep5 (raw)


title: LLC1 - cisplatine_rep2

description: c733_LLC1 - cisplatine_rep2 (raw)


title: LLC1 - cisplatine_rep1

description: c732_LLC1 - cisplatine_rep1 (raw)


title: AH316

description: Gene expression data


title: AM235

description: Gene expression data


title: AC270

description: Gene expression data


title: LLC1 - KW_rep1

description: c728_LLC1 - KW_rep1 (raw)


title: LLC1 - vehicle_vehicle_rep4

description: c751_LLC1 - vehicle_vehicle_rep4 (raw)


title: AL246

description: Gene expression data


title: LLC1 - vehicle_vehicle_rep5

description: c752_LLC1 - vehicle_vehicle_rep5 (raw)


title: LLC1 - KW_rep3

description: c753_LLC1 - KW_rep3 (raw)


title: AM194

description: Gene expression data


title: AA325

description: Gene expression data


title: AC209

description: Gene expression data


title: AM203

description: Gene expression data


title: AF264

description: Gene expression data


title: LLC1 - cisplatine_rep5

description: c759_LLC1 - cisplatine_rep5 (raw)


title: LLC1 - vehicle_vehicle_rep2

description: c749_LLC1 - vehicle_vehicle_rep2 (raw)


title: AA242

description: Gene expression data


title: LLC1 - vehicle_rep1

description: c726_LLC1 - vehicle_rep1 (raw)

value: vehicle


title: Kidney - Cisplatine + KW6002 rep6

description: c538_Kidney - Cisplatine + KW6002 rep6 (raw)

data_processing:
Image analysis and base calling were performed using RTA 2.7.3 and bcl2fastq 2.17.1.14.
Reads were mapped onto the mm10 assembly of Mus musculus genome using STAR [3] version 2.5.3a.
Gene expression quantication was performed from uniquely aligned reads using htseq-count version  0.6.1p1, with annotations from Ensembl version 90 and union mode.
Genome_build: mm10
Supplementary_files_format_and_content: Tab-separated text file with raw read counts per Ensembl Gene (ensembl v90).

source_name: Kidney

extract_protocol:
Total RNA was extracted by phenol/ chloroform and subsequently precipitated in isopropanol as described previously (Vomelováet al., 2009). Total RNAs from cultured renal cells were extracted usingRNeasy Mini kit (Qiagen) following manufacturer’s recommendations.
RNA-Seq libraries were generated from 300ng of total RNA using TruSeq Stranded mRNA LT Sample Preparation Kit (Illumina, San Diego, CA), according to manufacturer's instructions. Briefly, following purification with poly-T oligo attached magnetic beads, the mRNA was fragmented using divalent cations at 94oC for 2 minutes. The cleaved RNA fragments were copied into first strand cDNA using reverse transcriptase and random primers. Strand specificity was achieved by replacing dTTP with dUTP during second strand cDNA synthesis using DNA Polymerase I and RNase H. Following addition of a single 'A' base and subsequent ligation of the adapter on double stranded cDNA fragments, the products were purified and enriched with PCR (30 sec at 98oC; [10 sec at 98oC, 30 sec at 60oC, 30 sec at 72oC] x 12 cycles; 5 min at 72oC) to create the cDNA library. Surplus PCR primers were further removed by purification using AMPure XP beads (Beckman-Coulter, Villepinte, France) and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis.

value: Kidney

value: Cisplatine + KW6002


title: Kidney - Cisplatine + KW6002 rep5

description: c537_Kidney - Cisplatine + KW6002 rep5 (raw)


title: Kidney - Cisplatine + KW6002 rep4

description: c536_Kidney - Cisplatine + KW6002 rep4 (raw)


title: Kidney - Cisplatine + KW6002 rep3

description: c534_Kidney - Cisplatine + KW6002 rep3 (raw)


title: Kidney - Cisplatin rep2

description: c529_Kidney - Cisplatin rep2 (raw)

value: Cisplatin


title: Kidney - KW6002 rep4

description: c525_Kidney - KW6002 rep4 (raw)


title: Kidney - Cisplatin rep5

description: c532_Kidney - Cisplatin rep5 (raw)


title: Kidney - KW6002 rep5

description: c526_Kidney - KW6002 rep5 (raw)


title: Kidney - Vehicle rep4

description: c521_Kidney - Vehicle rep4 (raw)

value: Vehicle


title: LLC1 - cisplatine+KW_rep4

description: c760_LLC1 - cisplatine+KW_rep4 (raw)


title: Kidney - KW6002 rep2

description: c346_Kidney - KW6002 rep2 (raw)


title: LLC1 - vehicle_vehicle_rep3

description: c750_LLC1 - vehicle_vehicle_rep3 (raw)


title: Kidney - Cisplatine + KW6002 rep1

description: c524_Kidney - Cisplatine + KW6002 rep1 (raw)


title: Kidney - Vehicle rep2

description: c339_Kidney - Vehicle rep2 (raw)


title: Kidney - Vehicle rep3

description: c340_Kidney - Vehicle rep3 (raw)


title: Kidney - KW6002 rep6

description: c535_Kidney - KW6002 rep6 (raw)


title: Kidney - Cisplatin rep1

description: c528_Kidney - Cisplatin rep1 (raw)


title: Kidney - KW6002 rep3

description: c347_Kidney - KW6002 rep3 (raw)


title: Kidney - Vehicle rep5

description: c523_Kidney - Vehicle rep5 (raw)


title: Kidney - Vehicle rep1

description: c337_Kidney - Vehicle rep1 (raw)


title: 170322_RNA3B

status: Public on Aug 22 2022

city: Moscow

first: Ivan

middle: Nikolaevitch

last: Vlasov

address: Akademika Kurchatova square, 2

country: Russia

institute: National Research Center «Kurchatov Institute»

data_processing:
3 different pipelines were used in the process of data processing.
Fastq files for each sample were aligned to the GRCh38 genome (Gencode, release 37) using RSEM.
Ambiguous and low-quality bases were removed from the files obtained during the FASTQ sequencing process. Removal was done with AdapterRemoval V2. The trimmed files were aligned to the human reference genome GRCH38 and GRCH38.92 gene annotation using RSEM with rsem-prepare-reference and rsem-calculate-expression commands  and the -star option to also generate STAR indices.
Assembly: GRCH38
Supplementary files format and content: Processed data files are generated by RSEM and contain normalized and raw per gene counts.

source_name: transcriptome of blood of twin with parkinson 3 — replication 1

extract_protocol:
Total RNA isolation was conducted using a Direct-Zol RNA Mini-Prep Kit (ZymoResearch) according to the manufacturer's recommendations. The concentration of total RNA isolated was measured using a Quant-iT RNA BR Assay Kit and a Qubit fluorimeter (Invitrogen). RNA quality was monitored using an Experion automated electrophoresis system (Bio-Rad Laboratories).
Poly-a fraction of RNA was extracted from total RNA for RNA-seq. Sequencing libraries were prepared using NEBNext® mRNA Library Perp Reagent Set (NEB, USA). Sequencing was performed using HiSeq1500 (Illumina, USA), obtaining no less then 15 mln. reads of 50 length per library.

value: blood

value: Batch 2

tag: parkinson status

value: PD


title: Kidney - KW6002 rep1

description: c345_Kidney - KW6002 rep1 (raw)


title: 170322_RNA3K

source_name: transcriptome of blood of twin control 3 — replication 1

value: control


title: 170322_RNA2B

source_name: transcriptome of blood of twin with parkinson 2 — replication 1


title: 160322_RNA1K_4

source_name: transcriptome of blood of twin control 1 — replication 4

value: Batch 1


title: 160322_RNA1K_5

source_name: transcriptome of blood of twin control 1 — replication 5


title: Kidney - Cisplatine + KW6002 rep2

description: c533_Kidney - Cisplatine + KW6002 rep2 (raw)


title: 170322_RNA2K

source_name: transcriptome of blood of twin control 2 — replication 1


title: Kidney - Cisplatin rep3

description: c530_Kidney - Cisplatin rep3 (raw)


title: 160322_RNA1K_3

source_name: transcriptome of blood of twin control 1 — replication 3


title: 160322_RNA1K_6

source_name: transcriptome of blood of twin control 1 — replication 6


title: 160322_RNA1K_2

source_name: transcriptome of blood of twin control 1 — replication 2


title: Kidney - Cisplatin rep4

description: c531_Kidney - Cisplatin rep4 (raw)


title: 160322_RNA1B_5

source_name: transcriptome of blood of twin with parkinson 1 — replication 5


title: 160322_RNA1K_1

source_name: transcriptome of blood of twin control 1 — replication 1


title: 160322_RNA1B_2

source_name: transcriptome of blood of twin with parkinson 1 — replication 2


title: 160322_RNA1B_3

source_name: transcriptome of blood of twin with parkinson 1 — replication 3


title: 160322_RNA1B_4

source_name: transcriptome of blood of twin with parkinson 1 — replication 4


title: 160322_RNA1B_1

source_name: transcriptome of blood of twin with parkinson 1 — replication 1


title: 160322_RNA1B_6

source_name: transcriptome of blood of twin with parkinson 1 — replication 6


title: J187_6

status: Public on Jul 27 2022

city: Philadelphia

first: Jai-Yoon

last: Sul

address: 89 John Morgan

institute: University of Pennsylvania

data_processing:
Following sequencing, adapters, library barcode sequences, polyA tails and low quality bases (phred score < 20) were trimmed from all reads using a custom python script prior to alignment to the mouse genome (GENCODE GRCm38) using STAR (https://github.com/safisher/ngs/blob/master/ngs_STAR.sh).
Feature counts of high quality uniquely aligned reads were quantified using VERSE (modified HTSeq with hierarchical assignment: exon, mito, intron, intergenic).
The raw count matrix and the metadata can be found in raw_count_matrix.csv.gz.
Assembly: mm10
Supplementary files format and content: comma-separated values (CSV) file including the metadata and raw gene counts

source_name: No-response astrocyte

growth_protocol: Timed pregnant C57BL/6 mice were purchased from Charles River. All procedures were approved by the University of Pennsylvania’s Institutional Animal Care Committee. Ethanol-200 proof (Decon Labs #2716), was obtained from Decon Labs through the Alcohol Service Center at the University of Pennsylvania. Other chemicals used in the recording solution for Calcium imaging were purchased from Sigma.

extract_protocol:
Coverslips containing plated astrocytes were loaded with 10μg/ml Fluo-4 AM (#F14201, Thermo Fisher Scientific) mixed with 0.0001% pluronic acid (#P3000MP, Thermo Fisher Scientific) for 25min at room temperature then de-esterified for 10 mins with continual perfusion of external solution. Serum was removed 24 hours prior to experiments. External solution (Imaging saline, perfusion speed: 3ml/min) contained 140mM NaCl, 5.4mM KCl, 1mM MgCl2, 2mM CaCl2, 16mM glucose, 10mM HEPES, adjusted to pH 7.3. The calcium indicator-loaded coverslip was then transferred to an open bath chamber (RC-27, Warner Instruments) for imaging and sample collection. To minimize cell-to-cell fluorescence loading variation and animal and culture batch variations, the calcium signal in individual astrocytes was normalized against calcium signals induced by 100µM ATP at the end of each experiment. For single cell isolation, targeted single cells were identified based on real-time imaging results and collected with a pipet tip pulled from a glass microcapillary (#3450099, Thermo Fisher Scientific) controlled by a micromanipulator (Eppendorf, Transferman NK2). Immediately after isolating samples by gentle aspiration, samples were snap-frozen on an ethanol-dry ice mixture then briefly stored at -20oC prior to mRNA amplification
Single cells were collected in 5x first strand buffer (ThermoFisher, supplied with #1808004) and snap frozen prior to storage. Single cells were then thawed and amplified using modifications to the antisense RNA (aRNA) amplification procedure (80). Alterations included using Beckman Coulter Ampure XP beads (#A63881) for all cleanups after second strand reaction or the Beckman Coulter Agencourt RNAClean XP (#A63987) following the in vitro transcription reaction. Cleaned double stranded cDNA or cleaned aRNA was eluted in small volumes to allow direct input into the next step without the need for an ethanol precipitation or speed vacuum concentration step. Amplification was restricted to two rounds to ameliorate the shortening effect of random primed based approaches as well as the severity of 3’bias of polyA selection amplification approaches. Second round aRNA was sheared based on overall size distribution assessed using an Agilent Tapestation 2200, with the shearing time adjusted for longer average aRNA products, prior to input into the Illumina TruSeq mRNA stranded library construction procedure (#20020594, set A indexes #20020492, and set B indexes #20020493). A total of 50ng of aRNA from a single sample was input into library construction. All other steps after shearing were followed according to the manufacturer’s specifications. Following completion of library preparation, samples were multiplexed at 12 samples per lane and sequenced on a NextSeq 500 using the 150 cycle high output v2 kit (#FC-404-2002), allowing for paired end 2x75 base pair reads.

treatment_protocol: Primary cultured astrocytes were prepared from cortex of single C57BL/6 mouse pups (P0-P1) as described. In brief, the cerebral cortex was dissected from the brain and adherent meninges were removed prior to mincing and dissociation with 2.5% trypsin (20 mins, #15090-046, Gibco) by trituration. This single cell suspension was then passed through a 40 m cell strainer (#22363547, Fisher Scientific) to remove clumps. Cells were plated in 25 cm2 plastic flasks (#sial0639, Sigma) and cultured in Dulbecco’s modified Eagle’s medium (DMEM, #10-013, Corning) supplemented with (in mM) 25 glucose, 4 L-glutamine, 1 sodium pyruvate and 10% fetal bovine serum (#100-500, GemCell). Cultures were maintained at 37°C in a humidified 5% CO2 incubator. The media was replaced every 2-3 days. Prior to re-plating on glass coverslips coated with 0.1 mg/ml poly L-lysine (PLL, #P9155, Sigma), cultures were shaken (250rpm for 6hrs) and treated for 12hours with cytosine arabinoside (10M) to remove potentially contaminating cell types such as microglia.

tag: cell type

value: primary astrocyte culture

tag: strain

tag: response

value: No Response

tag: region

value: cortex


title: J186_5


title: J185_4


title: J175_18

source_name: M-type astrocyte

value: M-type


title: J183_1


title: J182_2


title: J176_25


title: J174_13


title: SAT_20

status: Public on Aug 01 2022

description: Transcriptome-wide gene expression data from the subcutaneous adipose tissue (SAT) of a high-fat diet (HFD)-induced obese mouse treated with LY2584702 (LY) tosylate (HFD+LY)

hyb_protocol: Sample was hybridized to Affymetrix Clariom S Mouse array using an Affymetrix GeneChip Hybridization Oven 645

source_name: subcutaneous adipose tissue

label_protocol: 100 ng of total RNA from each sample was processed and labelled according to manual GeneChip WT PLUS Reagent kit (P/N 703174 Rev. 2, Affymetrix Inc., Santa Clara, CA, USA)

extract_protocol: Total RNA was extracted following Qiagen protocol (RNeasy Mini Kit, Cat.#74104)

value: subcutaneous adipose tissue

tag: gender

value: male

tag: treatment

value: treated (HFD+LY)


title: J172_23


title: SAT_11

description: Transcriptome-wide gene expression data from the subcutaneous adipose tissue (SAT) of a high-fat diet (HFD)-induced obese mouse treated with vehicle (control HFD)

value: untreated (HFD)


title: J173_25


title: J166_25

source_name: T-type astrocyte

value: T-type


title: SAT_16


title: SAT_18


title: J165_23


title: LIV_14

description: Transcriptome-wide gene expression data from the liver (LIV) of a high-fat diet (HFD)-induced obese mouse treated with vehicle (control HFD)

source_name: liver

value: liver


title: LIV_20

description: Transcriptome-wide gene expression data from the liver (LIV) of a high-fat diet (HFD)-induced obese mouse treated with LY2584702 (LY) tosylate (HFD+LY)


title: LIV_8


title: J162_16


title: J161_9


title: J156_16

source_name: S-type astrocyte

value: S-type


title: J163_18


title: J155_9


title: LIV_16


title: SAT_8


title: LIV_11


title: IgG_cells

status: Public on Jun 22 2022

description: IgG cells

data_processing:
Libraries were sequenced on an Illumina HiSeq 4000 sequencer as paired-end 100 base reads. Image analysis and base calling were performed using RTA version 2.7.7 and bcl2fastq version 2.20.0.422.
Data were preprocessed with Cutadapt (Martin 2011) v1.18 to trim adapter sequences (Nextera Transposase Sequence) from 3’ end of reads. Cutadapt was used with the following parameters ‘-a CTGTCTCTTATA -A CTGTCTCTTATA -m 25:25’. Reads were mapped to Mus musculus genome (assembly mm10) using Bowtie2 (Langmead and Salzberg 2012) v2.2.8 with default parameters except for “–end-to-end –very-sensitive –no-mixed –no-discordant -I 10 -X 700”.
Assembly: mm10
Supplementary files format and content: BigWig files were generated using Homer (Heinz et al. 2010) makeUCSCfile v4.11.0 with default parameters except for “-norm 10e6 -fragLength 200”.

source_name: hippocampus

extract_protocol:
Tagmented DNA was extracted using MinElute PCR Purification Kit (Qiagen, # 28004)
Library preparation was conducted according to the protocol using Nextera primers (Illumina, #FC-131-2001) and post-PCR clean-up using SPRI bead slurry (Beckmann Coulter, #B23317).

value: hippocampus

value: isolated hippocampal cells

tag: chip antibody

value: Rabbit IgG (Diagenode #C15410206)


title: J150_3


title: LIV_18


title: J153_6


title: SAT_14


title: J154_8


title: IgG_neurons

description: IgG neurons

value: isolated hippocampal neurons


title: cells_water_H3K27ac_1

description: cells water H3K27ac

value: H3K27ac (Abcam, #ab4729)


title: J164_19


title: cells_caff_H3K27ac_2

description: cells caffeine H3K27ac


title: cells_caff_H3K27ac_1

description: cells caffeine H3K27ac


title: neurons_caff_H3K27me3_2

description: neurons caffeine H3K27me3


title: neurons_caff_H3K27ac_2

description: neurons caffeine H3K27ac


title: neurons_water_H3K27ac_2

description: neurons water H3K27ac


title: neurons_caff_H3K27me3_1

description: neurons caffeine H3K27me3


title: neurons_water_H3K27ac_1

description: neurons water H3K27ac


title: H3K9/14ac_HCCaffeine_rep2

description: H3K9K14ac caffeine

data_processing:
Image analysis and base calling were performed using RTA 2.7.3 and bcl2fastq 2.17.1.14.
Sequence reads were mapped to reference genome mm10 using Bowtie 1.0.0 with the following parameters -m 1 --strata --best -y -S -l 40 -p 2.
Genome_build: mm10
Supplementary_files_format_and_content: Wig files were generated using with an in-house script (Variable step, span=25, reads were elongated to 200b).

extract_protocol:
Freshly dissected tissue was chopped by a razor blade and rapidly incubated in 1.5 mL PBS containing 1% formaldehyde for 10 minutes at RT. To stop fixation, glycine was added (0.125 M final concentration). Dorsal hippocampi from 4 mice were pooled per sample. Tissue samples were then processed as described in Chatterjee et al. 2018, and sonicated using the Diagenode Bioruptor (30 s ON-30 s OFF at High Power x 35 cycles). Sonicated chromatin was centrifuged 10 min at 14000xg, the supernatant collected and diluted 1:10 in ChIP dilution buffer (0.01% SDS, 1.1% Triton X-100, 1.2 mM EDTA, 16.7 mM Tris-Cl, pH 8.1, 167 mM NaCl). A fraction of the supernatant (50 mL – 10%) from each sample was saved before immune-precipitation for ‘total input chromatin’. Supernatants were incubated overnight (4°C) with 1/1000 primary antibody H3K9/14ac (Diagenode C15410200) and H3K27ac (ab4729), followed by protein A Dynabeads (Invitrogen) for 2 hours at RT. After several washes (low salt, high salt, LiCl and TE buffers), the resulting DNA-protein complexes were eluted in 300 mL elution buffer (1% SDS, 0.1 M NaHCO3). The crosslinking was reversed (overnight at 65°C) and the DNA was purified with RNAse (30 min, 37°C), proteinase K (2 hours at 45°C). DNA from the immunoprecipitated and input samples was isolated using Diagenode MicroChIP DiaPure columns with 20 mL nuclease-free milliQ water in low binding tubes.
ChIP samples were purified using Agencourt AMPure XP beads (Beckman Coulter) and quantified with the Qubit (Invitrogen). ChIP-seq libraries were prepared from 10ng of double-stranded purified DNA using the MicroPlex Library Preparation kit v2 (C05010014, Diagenode s.a., Seraing, Belgium), according to manufacturer's instructions. In the first step, the DNA was repaired and yielded molecules with blunt ends. In the next step, stem-loop adaptors with blocked 5 prime ends were ligated to the 5 prime end of the genomic DNA, leaving a nick at the 3 prime end. The adaptors cannot ligate to each other and do not have single-strand tails, avoiding non-specific background. In the final step, the 3 prime ends of the genomic DNA were extended to complete library synthesis and Illumina compatible indexes were added through a PCR amplification (4 + 7 cycles). Amplified libraries were purified and size-selected using Agencourt AMPure XP beads (Beckman Coulter) to remove unincorporated primers and other reagents.

treatment_protocol: Two-three month-old mice were randomly assigned to the two following experimental groups: water (control group) and caffeine. The caffeine solution was kept in dark bottles thus protected from light and changed weekly. Treatment started at eight to nine weeks of age and lasted for two weeks. The chronic caffeine treatment in mice has been set in order to mimic the usual dose range of caffeine consumption in Humans. The selected caffeine dose of 0.3 g/L p.o., administered through drinking water at 0.3 g/L, has been previously shown to provide a significant benefit in neurodegenerative contexts (Arendash et al., 2006; Arendash et al., 2009; Laurent et al., 2014).

value: caffeine


title: H3K9/14ac_HCwater_rep1

description: H3K9K14ac water

value: water


title: H3K9/14ac_HCCaffeine_rep1

description: H3K9K14ac caffeine


title: H3K9/14ac_HCwater_rep2

description: H3K9K14ac water


title: H3K27ac_HCCafeine_rep1

description: H3K27ac caffeine

value: H3K27ac (Abcam, ab4729)


title: H3K27ac_HCCaffeine_rep2

description: H3K27ac caffeine


title: cells_water_H3K27ac_2

description: cells water H3K27ac


title: H3K27ac_HC0_rep2

description: H3K27ac water


title: H3K27ac_HC0_rep1

description: H3K27ac water


title: J151_5

value: primary astrocyte culture

value: cortex


title: Input_HCCafeine

description: input caffeine


title: neurons_water_H3K27me3_2

description: neurons water H3K27me3


title: Input_HCwater

description: input water


title: HD36_MWM_water

data_processing:
Image analysis and base calling were performed using RTA 2.7.3 and bcl2fastq 2.17.1.14.
Reads were mapped onto the mm10 assembly of Mus musculus genome using STAR [3] version 2.5.3a.
Gene expression quantication was performed from uniquely aligned reads using htseq-count version  0.6.1p1, with annotations from Ensembl version 90 and union mode.
Genome_build: mm10
Supplementary_files_format_and_content: Tab-separated text file with raw read counts per Ensembl Gene.

source_name: Hippocampus

extract_protocol:
Total RNA was extracted from dorsal hippocampal tissues using TRIzol reagent (Invitrogen) (n=4/group). Freshly dissected tissue was chopped, homogenized in 300 μL of TRIzol reagent, and frozen (20 minutes at -80°C), followed by 3-minutes centrifugation at 14000xg before chloroform/isoamyl extraction. The supernatant was used to precipitate RNA using isopropanol and RNase-free glycogen (30 minutes at 4°C). The pellet was washed once with 70% ethanol and resuspended in Milli-Q water. A new RNA precipitation was performed with 100% ethanol and 3 M NaOAC (overnight at -20°C). After two further 70% ethanol washes, pellet was air-dried and resuspended in 30 μL nuclease-free Milli-Q water, heated 6 minutes at 50°C, and RNA quantification was performed.
RNA-Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded mRNA LT Sample Preparation Kit (Illumina, San Diego, CA), according to manufacturer's instructions. Briefly, following purification with poly-T oligo attached magnetic beads, the mRNA was fragmented using divalent cations at 94oC for 2 minutes. The cleaved RNA fragments were copied into first strand cDNA using reverse transcriptase and random primers. Strand specificity was achieved by replacing dTTP with dUTP during second strand cDNA synthesis using DNA Polymerase I and RNase H. Following addition of a single 'A' base and subsequent ligation of the adapter on double stranded cDNA fragments, the products were purified and enriched with PCR (30 sec at 98oC; [10 sec at 98oC, 30 sec at 60oC, 30 sec at 72oC] x 12 cycles; 5 min at 72oC) to create the cDNA library. Surplus PCR primers were further removed by purification using AMPure XP beads (Beckman-Coulter, Villepinte, France) and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis.

value: dorsal hippocampus

tag: group

value: Learning (water)


title: neurons_water_H3K27me3_1

description: neurons water H3K27me3


title: HD20_HC_water

value: Home cage (water)


title: HD11_HC_water


title: HD14_HC_Caffeine

value: Home cage (caffeine)


title: HD40_MWM_Caffeine

value: Learning (caffeine)


title: neurons_caff_H3K27ac_1

description: neurons caffeine H3K27ac

tag: condition

value: ca


title: HD30_MWM_Caffeine


title: HD33_MWM_water


title: HD27_MWM_Caffeine


title: HD22_MWM_Caffeine


title: HD24_MWM_water


title: HD6_HC_water


title: HD17_HC_Caffeine


title: HD38_MWM_water


title: HD2_HC_Caffeine


title: HEK293T, H3K27ac, HBG1, dCas9-CBP

status: Public on Jun 21 2022

city: Houston

first: Isaac

middle: B

last: Hilton

state: TX

address: 6500 Main St

department: Bioengineering

institute: Rice University

data_processing:
Basecalls were performed using bcl2fastq (v2.17)
Reads were trimmed using Trim Galore (v0.6.6)
Aligned to the hg38 reference genome using Bowtie2 (v7.3.0)
Aligned reads were deduplicated using Picard MarkDuplicates (v2.23.8)
Deduplicated alignments were converted into a bedGraph file using bedtools (v2.30.0)
BedGraph files from biological replicates were then merged using bedops (v2.4.39)
Assembly: hg38
Supplementary files format and content: bedGraph (except for IgG control)
Library strategy: CUT&RUN-Seq

growth_protocol: HEK293T cells (ATCC, CRL-11268) were cultured in Dulbecco’s modified Eagle’s medium (DMEM; Gibco, 31-053-028) supplemented with 10% FBS (Sigma, F2442) and 1% penicillin/streptomycin (Gibco, 15140). Cell lines were maintained at 37 °C and 5% CO2.

extract_protocol:
CUT&RUN assays were carried out using the Epicypher CUTANA ChIC/CUT&RUN Kit (Epicypher 14-1048). Briefly, 5.0e5 transfected cells were harvested, immobilized on concanavalin A beads, and permeabilized in 0.01% digitonin cell permeabilization buffer. The cell-bead conjugate mixture was then divided equally into 2 aliquots and incubated in 50ul antibody buffer with either 0.5µg of anti-H3K27ac antibody (Abcam, 4279) or 0.5µg of control rabbit IgG antibody (Epicypher, 14-1048) overnight at 4°C. After washing the beads, pAG-MNase was added to the immobilized cells and the solution was incubated for 2 hours at 4°C to digest and release chromatin DNA
Libraries were constructed using NEBNext® Ultra™ II DNA Library Prep Kit for Illumina® kit (NEB, E7645) by Azenta Life Sciences

tag: cell line

value: Human embryonic kidney


title: HEK293T, H3K27ac, HS2, dCas9-VPR


title: HEK293T, H3K27ac, HS2, dCas9-CBP