title: HEK293 Fast Pol II Nanopore

city: Hanover

first: Mareike

last: Polenkowski

address: Carl-Neuberg-Straße 1

department: Cell Biochemistry

institute: Medical School Hannover

data_processing:
Nanopore: Basecalls performed using Guppy version 3.3.3+fa743a6
Nanopore: Mapping using minimap2
Nanopore: Trimming of long read sequencing data using NanoFilt version 2.5.0
ChIp-Seq: FASTQ files were generated by CASAVA (v1.8.2). Galaxy workflow for RNA-Seq (www.usegalaxy.org) was used for subsequent data analysis. Reads were mapped to the human reference genome (GRCh38) using Bowtie2 (Galaxy Version 2.3.4.1). Metagene analysis was performed using deepTools 2.0
TT-Seq: Raw reads were mapped to the human reference genome (GRCh38) using Bowtie2 (Galaxy Version 2.3.4.1). Read coverage of individual gene was normalized and visualized using Seqmonk. To measure the progression of Pol II molecules into the gene body we applied a pipeline described by Gregersen et al. (33) for calling RNA Pol II transcription wave peak positions and elongation rates from DRB/TT-seq time-series data using R. Metagene analysis was performed using deepTools 2.0.
Assembly: GRCh38
Supplementary files format and content: For the identification of 3’cleavage sites the tool NanoFilt (Filtering and trimming of long read sequencing data) was used to trim the mRNA sequences from 5’end to a uniform length so that we keep always the last 200 nucleotides (python get_read_ends.py --bases_from_end 200 reads.fastq.gz | gzip > last_200_bp.fastq.gz). Trimmed reads were aligned to the GRCh38 human genome reference using minimap2. The quantification of 3’ cleavage was perform with Seqmonk using the human polyA database as reference. Genes that contain more than two annotated polyadenylation sites (PAS) and more than 5 reads at distal PAS sites were selected. The ratios between proximal- and distal cleavage site were calculated.

molecule: cytoplasmic RNA

source_name: kidney

growth_protocol: HEK293 cells were grown in Dulbecco´s modified Eagle's medium supplemented with 10% (v/v) FCS. HEK293 cells expressing Dox inducible mutant RNA Pol II that contain additional mutation for amanitine resistant (Fond et al ) were maintained in DMEM supplemented with 10% FBS, 200 μg/mL hygromycin B, 6.5 μg/mL blasticidin, and penicillin/streptomycin. Experiments using cells expressing Pol II mutants were performed after induction with 2.0 μg/mL doxycycline for 12–16 h and treatment with 2.5 μg/mL α-amanitin for a further 42 h, at which time all cell lines were viable, and endogenous Pol II was inactive.

extract_protocol:
Cytoplasmic RNA of HEK293 cells were achieved by RLN (Cyto-Buffer) and the Reliaprep™ miRNA Cell and Tissue Miniprep System (Promega). mRNAs were isolated by the NEBNext® Poly(A) mRNA Magnetic Isolation Module (E7490) with use of 30 µg RNA.
RNA libraries were prepared for sequencing using standard Nanopore protocols for Direct RNA sequencing (SQK-RNA002).

treatment_protocol: Infection of HEK293 cells with lentiviruses carrying either shCtrl or shTHOC5. The treatment was applied for 4 days. Transfection with siCtrl, siTHOC2, siTHOC6 or siCDK12 for 3 days.

value: kidney

value: Human embryonic kidney 293 cells

value: lentiviral infection with shCtrl or shTHOC5, 1-3 days, then doxycycline for 12-16h and α-amanitin for 42h


title: HEK293 siCDK12 Nanopore

status: Public on Dec 16 2022

type: SRA

country: Germany

description: 626 THOC5 dependent genes_cytoplasmic expression.xlsx

library_source: transcriptomic

organism: Homo sapiens

tag: tissue

tag: cell line

tag: treatment

value: transfection with siCDK12, 3 days


title: HEK293 Slow Pol II Nanopore


title: HEK293 siTHOC2 Nanopore

value: transfection with siTHOC2, 3 days


title: HEK293 shTHOC5-2 Nanopore

description:
626 THOC5 dependent genes_cytoplasmic expression.xlsx
ratios of 213 THOC5 independent genes.xlsx
ratios of 626 THOC5 dependent genes.xlsx

value: lentiviral infection carrying shTHOC5, 4 days


title: HEK293 siTHOC6 Nanopore

value: transfection with siTHOC6, 3 days


title: HEK293 siCtrl Nanopore

value: transfection with siCtrl, 3 days


title: HEK293 shTHOC5-1-repeat Nanopore


title: HEK293 shCtrl-repeat Nanopore

value: lentiviral infection carrying shCtrl, 4 days


title: neurospheres, CBPf/f

status: Public on Dec 17 2022

city: San Juan de Alicante

first: Ángel

last: Márquez Galera

address: Av. Santiago Ramón y Cajal, s/n

country: Spain

institute: Instituto de Neurociencias

library_source: transcriptomic single cell

data_processing:
Libraries were sequenced together in a unique cell flow on an Illumina HiSeq2500 sequencer to obtain 75 bp paired-end reads following sequencer manufacturer’s instructions sequencer, with an average sequencing depth of an average of 290 million reads per sample. Quality control was performed using FastQC v0.11.9. Sequenced samples were processed using the Cell Ranger v3.1 pipeline (10X Genomics) and aligned to the GRCm38 (mm10) mouse reference genome customized to count reads in exons and introns (pre-mRNA) (gene annotation version 94).
Bioinformatic analysis were performed as described previously in (Lipinski et al., 2020). Single-nucleus RNA-seq data were pre-processed and further analyzed in R using Seurat v2.3.4 (Butler et al., 2018; Stuart et al., 2019) using the following filter parameters: genes, nCell < 5; cells, nGene < 200. Barcodes with total unique molecular identifier (UMI) count > 10% of the 99th percentile of the expected recovered cells were selected for further analysis. We retrieved 4,884 (control), 3,017 (Crebbpf/f+Cre) and 4,648 (Ep300f/f+Cre) high nuclei per sample. Mean reads per nucleus were 62,810 (control), 99,066 (Crebbpf/f+Cre) and 65,928 (Ep300f/f+Cre). Data were then normalized using global-scaling normalization (method: LogNormalize, scale.factor = 10,000). To identify major cell populations in the differentiated cultures, datasets were analyzed separately.
Highly variable genes (HVGs) were detected using FindVariableGenes function with default parameters. Then, normalized counts on HVGs were scaled and centered using ScaleData function with default parameters. Principal component analysis (PCA) was performed over the first ranked 1000 HVGs. Plots of the two principal components of the PCA, where cells were colored by dataset, excluded the presence of batch effects. Cluster detection was carried out with Louvain algorithm using 20 first PCA dimensions at resolution = 0.6 (the default and the optimal according to cell number, data dispersion, and co-expression of previously reported cell markers). Visualization and embedding were performed using t t-Distributed Stochastic Neighbor Embedding (tSNE)  (Laurens van der Maaten, 2008) and Uniform Manifold Approximation and Projection (UMAP) (McInnes et al., 2018) over PCA using the 20 first PCA dimensions.
After characterizing the cell heterogeneity of each condition separately, data from the control, CBP-ablated, and p300-ablated conditions were merged into a single data set. Using the first 1,000 HVGs from each sample, a total of 1,421 unique HVGs detected in the 3 conditions were obtained. Normalized counts on merged HVGs were scaled and centered using ScaleData function with default parameters and PCA was performed over them. Plots of the two principal components of the PCA, where cells were colored by dataset, excluded the presence of batch effects. Subsequent analyses repeated the same strategy and the same clustering and visualization parameters as in the individual analyses.  FindAllMarkers function with default parameters was used to identify gene markers for each cluster and to assign the cell-type identity. Populations were identified combining the clustering results over the merged dataset with clustering information obtained from the datasets separately, together with co-expression of population markers and information from external scRNA-seq datasets from the Allen Brain Map portal (Yao et al., 2021) and the Atlas of the Developing Mouse Brain (La Manno et al., 2021). Twelve clusters were unbiasedly identified but three of them were closely distributed and manually fused since they presented minor expression differences. UMAP plots of gene expression show normalized count (UMIs) per nucleus. Clustering was performed on merged dataset from control, CBP-ablated and p300-ablated conditions and populations were identified combining these results with clustering information obtained in the datasets separately, together with co-expression of population markers. FindAllMarkers function with default parameters was used to identify gene markers for each cluster and to assign cell-type identity to clusters based on scRNA-seq dataset of the Allen Brain Map portal (Mouse Whole Cortex and Hippocampus SMART-seq (2019) with 10x-SMART-seq taxonomy (2020).
Differential expression analysis (DEA) was used to identify population gene effects between the different conditions. For DEA, the nuclei of each population were compared against all the other nuclei in the merged dataset using Wilcoxon Rank Sum test on normalized counts with the following parameters (logfc.threshold = 0.25; min.pct = 0.25). Gene Ontology (GO) functional enrichment analyses for Biological Process (BP) were performed using clusterProfiler (v3.15.2). All enriched terms were considered significant at adjusted P values by BH cutoff < 0.05. The reference gene set used to perform the analysis was C5 (GO Biological Process) collection from the Molecular Signatures Database (MSigDB) (v6.2).
Assembly: GRCm38 (mm10)
Supplementary files format and content: Cell Ranger output files (barcodes.tsv, features.tsv, matrix.mtx)

molecule: nuclear RNA

organism: Mus musculus

source_name: Brain

extract_protocol:
For the single nucleus RNA-Seq experiment, a pool of ~15 differentiated neurosphere cultures from control, CBPf/f+Cre and p300f/f+Cre at 15DIV were used. Differentiation medium II was removed and wells were washed with cold HBSS. Cold Lysis Buffer (10 mM Tris-HCl, 10 mM NaCl, 3 mM MgCl2, and 0.1% NonidetTM P40 Substitute in Nuclease-Free Water) was added (500 µl) immediately to every well, gently pipetted until cells were completely suspended and kept on ice for 5 min to lyse cells. The nuclei suspension from the different samples (control, CBPf/f+Cre and P300f/f+Cre) was transferred to a 15ml tube and centrifuged at 500 rcf for 5 min at 4°C. Supernatant were removed and nuclei pellet was resuspended in 1 ml of Nuclei Wash and Resuspension Buffer (1X PBS with 1.0% BSA and 0.2 U/μl RNase Inhibitor). Centrifugation and nuclei wash steps were repeated twice.
Nuclei suspension was sorted on FACS Aria III (BD Biosciences) at 4°C and immediately proceed to 10x Genomics single cell protocol. 16,000 nuclei per sample (pool of ~15 differentiated neurosphere cultures) were loaded into single cell Chromium Chip B and Chromium Controller was run to generate partitioning thousands of nuclei into nanoliter-scale Gel Beads-in-emulsion (GEMs) and immediately reversed transcribed, where all generated full-length cDNA (from poly-adenylated mRNA) share a common 10x Barcode. Post-GEM-RT cleanup, cDNA amplification and 3ʹ Gene Expression Library Construction were performed with Chromium Single Cell 3’ Library & Gel Bead Kit v3. Quality control of cDNA and libraries were performed with Agilent Bioanalyzer in a High Sensitivity chip.

value: Brain

tag: cell type

tag: genotype

value: CBPf/f

value: tamoxifen


title: HEK293 shCtrl Nanopore

description:
cytoplasmic RNA
3002_Proximal_vs_Distal_ratio.xlsx
Cytoplasmic_transcript_abundance_of_1113_THOC5_target_genes.xlsx

data_processing:
Nanopore: Basecalls performed using Guppy version 3.3.3+fa743a6.
Nanopore: Mapping using minimap2.
Nanopore: Trimming of long read sequencing data using NanoFilt version 2.5.0.
ChIP-Seq: FASTQ files were generated by CASAVA (v1.8.2). Galaxy workflow for RNA-Seq (www.usegalaxy.org) was used for subsequent data analysis. Reads were mapped to the human reference genome (GRCh38) using Bowtie2 (Galaxy Version 2.3.4.1). Metagene analysis was performed using deepTools 2.0.
Genome_build: GRCh38
Supplementary_files_format_and_content: For the identification of 3’cleavage sites the tool NanoFilt (Filtering and trimming of long read sequencing data) was used to trim the mRNA sequences from 5’end to a uniform length so that we keep always the last 200 nucleotides (python get_read_ends.py --bases_from_end 200 reads.fastq.gz | gzip > last_200_bp.fastq.gz). Trimmed reads were aligned to the GRCh38 human genome reference using minimap2. The quantification of 3’ cleavage was perform with Seqmonk using the human polyA database as reference. Genes that contain more than two annotated polyadenylation sites (PAS) and more than 5 reads at distal PAS sites were selected. The ratios between proximal- and distal cleavage site were calculated.
Supplementary_files_format_and_content: bigWig files

molecule: total RNA

source_name: Human embryonic kidney 293 cells

growth_protocol: HEK293 cells were grown in Dulbecco´s modified Eagle's medium supplemented with 10% (v/v) FCS. HEK293 cells expressing Dox inducible mutant RNA Pol II that contain additional mutation for amanitine resistant (Fond et al.) were maintained in DMEM supplemented with 10% FBS, 200 μg/mL hygromycin B, 6.5 μg/mL blasticidin, and penicillin/streptomycin. Experiments using cells expressing Pol II mutants were performed after induction with 2.0 μg/mL doxycycline for 12–16 h and treatment with 2.5 μg/mL α-amanitin for a further 42 h, at which time all cell lines were viable, and endogenous Pol II was inactive.

treatment_protocol: Infection of HEK293 cells with lentiviruses carrying either shCr or shTHOC5. The treatment was applied for 4 days.

value: embryonic kidney cell line

value: lentiviral infection carrying shCtrl

tag: treatment duration

value: 4 days


title: Control base rep2

city: Louisville

first: Jae Yeon

last: Hwang

state: Kentucky

address: 505 S. Hancock Street

department: Division of Medical Oncology & Hematology

country: USA

institute: University of Louisville

data_processing:
HiSeq Control Software v3.3 for system control and base calling through Real Time Analysis v2.5.2 (Illumina)
Sequencing reads in Fastq files with three replicates of each sample group were aligned to Mus musculus genome (assembly GRCm38) with STAR aligner (version 2.5.2b).
Read counts and fragments per kilobase of transcript per million mapped reads (FPKM) were calculated based on the mouse GRCm38 Ensembl Release 82 gene annotation. Cuffdiff (version 2.2.1) was used for searching differentially expressed genes (DEGs) setting pairwise comparison for all sample groups at once.
Assembly: mm10
Supplementary files format and content: tab-delimited text file includes read counts, FPKMs, averaged FPKMs for each sample group

source_name: cochlea

extract_protocol:
cochleae were dissected from 4-week-old control and Fst cKO mutant mice and divided into three regions along the tonotopic axis, base, middle, and apex. Each cochlear region from each genotype was pooled together and RNAs were extracted.
The RNAs were divided into three technical replicates and were prepared for sequencing using the TruSeq RNA Sample Prep Kit v2 with Ribo-Zero (Illumina)

value: cochlea

value: WT

value: untreated

tag: age

tag: region

value: base


title: SINV in 293T

city: Wuhan

first: Yang

last: Qiu

state: Hubei

address: No. 44 Xiaohongshan Central District, Wuchang District,

country: China

institute: Wuhan institue of virology, CAS

description:
SINV (GenBank: NC_001547.1)
small RNA

data_processing:
Fo small RNAs, libraries of small RNAs were constructed using TruSeq Small RNA Library Preparation Kits (Illumina) according to manufacturer’s protocols, and then sequenced by Illumina HiSeq 2000 at Bioacme (Wuhan, China). Quality control and adapter trimming were performed using Trim Galore (v0.6.4) with default parameters, and reads length between 17 and 70 were used for subsequent analyses. The sequenced reads were mapped to the human genome (release hg19), mouse genome (release GRCm38), and mosquito genome (AaeGL5.0) by using Bowite 2.2.5, and the cellular miRNA expression level was determined on the basis of the number of reads, which were 100% identical to the annotated miRNA sequences in miRBase version 22 (https://www.mirbase.org/ftp/CURRENT/genomes/dme.gff3). The vsRNA reads that cannot be mapped to the host genome were then mapped to the viral genome allowing 2 nt mismatch. The vsRNA reads were counted and normalized by the number of total vsRNA reads (counts per million of total vsRNA reads) to show the size distribution and abundance of vsRNAs in different size, and 21-23 nt size vsRNA reads mapping to the viral genome were counted to show the distribution across the viral genome. The vsRNA Reads mapped to (+)-strand and (-)-strand of the viral genome were intersected using BedTools (v2.29.2).
For total RNAs, libraries of total RNAs were constructed using MGIEasy RNA Library Preparation Kits (BGI) according to manufacturer’s protocols, and then sequenced by MGISEQ2000 (BGI) at Wuhan Institute of Virology, CAS. Quality control and adapter trimming were performed using Trim Galore (v0.6.4) with default parameter. Then all reads were mapped to the human genome (release hg19) using Hisat2. DEseq2 (v1.34.0) was used to detect differentially expressed genes.
Assembly: Human genome (release hg19), mouse genome (release GRCm38), and mosquito genome (AaeGL5.0)

source_name: Human  HEK293T cells

extract_protocol: TruSeq Small RNA Library Preparation Kits (Illumina) (for small RNAs); MGIEasy RNA Library Preparation Kits (BGI) (for total RNAs)

value: HEK293T cells

value: Sindbis virus


title: SFV in 293T AGO2-KO #3

description:
SFV4 strain (GenBank: KP699763.1)
small RNA

value: Semliki Forest virus


title: SFV in 293T AGO2-KO #1


title: SFV in 293T AGO2-KO #2


title: SFV in 293T NoDice #2


title: SFV in 293T NoDice #1


title: SFV in 293T #3


title: SFV in 293T #2


title: SFV in 293T #1


title: Papillary RCC - RNA-seq [RT0642_RNA]

city: Boston

first: Elio

last: Adib

address: 20 Shattuck Street

institute: Brigham and Women's Hospital

description:
RT0642
RCC_RNAseq.tsv

data_processing:
FASTQ files were processed using the VIPER workflow.
Read alignment to human genome build hg19 was performed with STAR.
Cufflinks was used to assemble transcript-level expression data from filtered alignments.
Assembly: hg19
Supplementary files format and content: tab-delimited text file includes a matrix with FPKM values for each Sample

source_name: RCC tumor

extract_protocol:
RNA was extracted using the Qiagen RNeasy Mini Kit (Cat No./ID: 74104) as recommended, from frozen tumor samples adjacent to those samples used for ChIP.
RNA-seq libraries were constructed from 1 μg total RNA using the Illumina TruSeq Stranded mRNA LT Sample Prep Kit according to the manufacturer’s protocol.

value: RCC tumor

tag: tissue type

value: Papillary RCC


title: Papillary RCC - RNA-seq [RT1205_RNA]

description:
RT1205
RCC_RNAseq.tsv


title: Clear cell RCC - RNA-seq [RT0638_RNA]

description:
RT0638
RCC_RNAseq.tsv

value: Clear cell RCC


title: Clear cell RCC - RNA-seq [RT970_RNA]

description:
RT970
RCC_RNAseq.tsv


title: Papillary RCC - RNA-seq [RT1240_RNA]

description:
RT1240
RCC_RNAseq.tsv


title: Papillary RCC - RNA-seq [RT1184_RNA]

description:
RT1184
RCC_RNAseq.tsv


title: Clear cell RCC - RNA-seq [RT1097_RNA]

description:
RT1097
RCC_RNAseq.tsv


title: Papillary RCC - RNA-seq [RT1265_RNA]

description:
RT1265
RCC_RNAseq.tsv


title: HEK293 shTHOC5-1 Nanopore

value: lentiviral infection carrying shTHOC5


title: Clear cell RCC - RNA-seq [S22741_RNA]

description:
S22741
RCC_RNAseq.tsv


title: SFV in 293T NoDice #3


title: Clear cell RCC - RNA-seq [S20449_RNA]

description:
S20449
RCC_RNAseq.tsv


title: Clear cell RCC - RNA-seq [RT957_RNA]

description:
RT957
RCC_RNAseq.tsv


title: Clear cell RCC - RNA-seq [RT1043_RNA]

description:
RT1043
RCC_RNAseq.tsv


title: Clear cell RCC - RNA-seq [S21381_RNA]

description:
S21381
RCC_RNAseq.tsv


title: Chromophobe RCC - RNA-seq [RT0356_RNA]

description:
RT0356
RCC_RNAseq.tsv

value: Chromophobe RCC


title: Clear cell RCC - RNA-seq [RT1046_RNA]

description:
RT1046
RCC_RNAseq.tsv


title: Clear cell RCC - RNA-seq [RT1131_RNA]

description:
RT1131
RCC_RNAseq.tsv


title: Chromophobe RCC - RNA-seq [RT0301_RNA]

description:
RT0301
RCC_RNAseq.tsv


title: Chromophobe RCC - RNA-seq [RT1039_RNA]

description:
RT1039
RCC_RNAseq.tsv


title: Chromophobe RCC - RNA-seq [RT1002_RNA]

description:
RT1002
RCC_RNAseq.tsv


title: Chromophobe RCC - RNA-seq [RT1056_RNA]

description:
RT1056
RCC_RNAseq.tsv


title: Chromophobe RCC - RNA-seq [RT1059_RNA]

description:
RT1059
RCC_RNAseq.tsv


title: Chromophobe RCC - RNA-seq [RT929_RNA]

description:
RT929
RCC_RNAseq.tsv


title: Chromophobe RCC - RNA-seq [RT1152_RNA]

description:
RT1152
RCC_RNAseq.tsv


title: Chromophobe RCC - RNA-seq [RT1143_RNA]

description:
RT1143
RCC_RNAseq.tsv


title: Chromophobe RCC - RNA-seq [RT991_RNA]

description:
RT991
RCC_RNAseq.tsv


title: Chromophobe RCC - RNA-seq [RT1149_RNA]

description:
RT1149
RCC_RNAseq.tsv


title: Papillary RCC - RNA-seq [RT998_RNA]

description:
RT998
RCC_RNAseq.tsv


title: Clear cell RCC - RNA-seq [RT1047_RNA]

description:
RT1047
RCC_RNAseq.tsv


title: Err2, Rep 6

city: Tübingen

first: Vikas

last: Bansal

state: BW

address: Otfried Müller Strasse 23

institute: German Center for Neurodegenerative Diseases (DZNE)-Tübingen

data_processing:
RNA-sequencing quality was evaluated by utilizing raw reads using the FastQC quality control tool version 0.10.1
Bowtie2 v2.0.2 using RSEM version 1.2.29 with default parameters was utilized to align sequence reads (single-end 50 bp) to chicken reference genome (Galgal5)
Prior to indexing, GFP, ERR2, Dlk-1, VP16 and IRES sequences and annotations were added to the reference genome (FASTA file) and annotations (GTF file). Ensembl annotations (version 86.5) with rsem-prepare-reference from RSEM software was used to index chicken reference genome
Furthermore, sequence alignment of sequence reads and gene quantity was obtained through the use of rsem-calculate-expression. Rsem-calculate-expression resulted in sequence read count and TPM value (transcripts per million) for individual genes.
DESeq2 package was used to carry out differential expression analysis. Finally, genes with less than 5 reads (baseMean) were filtered, while genes with an adjusted p-value < 0.05 were classified as differentially expressed.
Assembly: Galgal5
Supplementary files format and content: Tab-delimited excel files (xlsx) include DESeq2 normalized values for each Sample

organism: Gallus gallus

source_name: Spinal motor column

growth_protocol: Lohmann LSL strain fertilized chicken eggs were incubated at 37.8°C and 80% humidity until E2.7-3.0 (HH St. 14-18). The embryos were then extracted 5 ml of their albumin using a syringe and partially removed of their eggshell to expose the embryo. The DNA constructs were injected into the central canal of the neural tube.

extract_protocol:
RNA was isolated using Tri-Reagent (Sigma-Aldrich Chemie GmbH, Taufkirchen, Germany) and Phenol-Chloroform extraction according to the manufacturer’s protocol.
RNA quality was assessed using Nanodrop 2000 (Thermo Fisher Scientific, GmbH) and RNA integrity number (RIN) was evaluated by using the Agilent 2100 Bioanalyzer (Agilent Technologies, Inc., USA). RNA was reverse transcribed to cDNA using Transcriptor High Fidelity cDNA synthesis kit (Roche Diagnostics Deutschland GmbH, Mannheim, Germany)
RNA-Seq libraries were obtained using TruSeq RNA Sample Preparation v2 kit (Illumina, Inc., San Diego, USA). To analyze the library quality, Agilent 2100 Bioanalyzer (Agilent Technologies, Inc., Santa Clara, USA) was used and the concentration was measured by a QubitR dsDNA HS Assay kit (Thermo Fisher Scientific Inc., Waltham, USA). The concentration was adjusted to 2 nM prior to sequencing (50 bp)

treatment_protocol: Chick embryos were electroporated with either eGFP (CMV-eGFP), or mouse Err2 (CMV-VP16-ERR2-eGFP), or mouse Dlk1 (pCAGGS-DLK1-IRES-eGFP) DNA constructs and spinal cord columns were identified by the eGFP signal, dissected and collected at E12.5 (HH St. 38-39) for the RNA sequencing studies.

value: Spinal motor column

value: motor neurons

value: ERR2 overexpression

value: in ovo electroporation

tag: time

value: 10 days


title: eGFP, Rep 4

value: eGFP overexpression


title: Err2, Rep 3


title: Err2, Rep 5


title: eGFP, Rep 3


title: Err2, Rep 2


title: eGFP, Rep 1


title: Err2, Rep 4


title: eGFP, Rep 5


title: eGFP, Rep 2


title: Dlk1, Rep 2

value: DLK1 overexpression


title: Dlk1, Rep 4


title: Dlk1, Rep 5


title: Dlk1, Rep 8


title: Dlk1, Rep 1


title: Err2, Rep 1


title: Dlk1, Rep 3


title: Dlk1, Rep 7


title: Dlk1, Rep 6


title: E12_LV_Tbx5_Mut_1

city: Cincinnati

first: Jeffrey

middle: D

last: Steimle

email: jeffrey.steimle@cchmc.org, jeffrey.steimle@bcm.edu, jsteimle@uchicago.edu, jsteimle@alumni.nd.edu

state: OH

address: 240 Albert Sabin Way

department: Molecular Cardiovascular Biology

institute: Cincinnati Children's Medical Hospital Center

data_processing:
Reads were aligned to the mouse genome (mm10, retrieved from UCSC May 23, 2012) using TopHat2 (version 2.1.1)
Reads were filtered to remove low-quality reads (MAPQ<10), duplicates, and unmapped reads using BamTools (version 2.4.1)
Post-alignment, post-filtered reads were assigned to transcripts using featureCounts from the Rsubread package v1.22.2 in R v3.3.2 (R Core Team 2016)
Differential expression testing was performed using edgeR v3.16.5 and limma v3.30.13. Low level genes were removed within each condition using median log-transformed counts per gene per million mapped reads (cpm) cutoff of 0. A generalized linear model (GLM) framework was used to test for differential expressed genes between mutants and controls.
Genome_build: mm10
Supplementary_files_format_and_content: The file 'counts_summarized.txt' contains transcript counts identified by featureCounts for each of the 7 samples (tab-deliminated; samples by column). The file 'E12_LV_TBX5CKO_significantGenes.txt' contains the genes that were significantly dysregulated between control and mutant samples.

molecule: polyA RNA

source_name: E12.5 left ventricle

extract_protocol:
At E12.5, embryos were dissected from the mother and the left ventricle microdissected. Total RNA was extracted using a combination of TRIzol and the RNeasy kit.
Isolated RNA from the left ventricle underwent mRNA selection using an oligo-dT pulldown, library preparation using the Illumina TruSeq RNA Sample prep kit v2 (Part# RS-122-2001), quantified and pooled in equimolar amounts using the Agilent Bio-analyzer 2100, and 50bp single-ended RNA-sequencing on the HiSeq2500 (Invitrogen) to a depth of approximately 26 million reads at the University of Chicago Genomics Facility

treatment_protocol: The pregnant female was treated with tamoxifen when the embryos were at E11.5.

tag: developmental stage

value: left ventricle


title: E12_LV_Tbx5_Ctrl_2


title: E12_LV_Tbx5_Ctrl_4


title: E12_LV_Tbx5_Ctrl_5


title: E12_LV_Tbx5_Mut_2


title: THP-1-D

city: Cambridge

first: Fadi

middle: John

last: Najm

state: Massachusetts

address: 415 Main Street 6043

department: Epigenomics

institute: Broad Institute

description:
Column name in the processed data file: THP-1-D
300k_conditions.txt
300k_reference.txt
300k_counts.txt

library_source: genomic

data_processing:
Data were processed with PoolQ to split sample reads by Illumina barcode (conditions) and count barcode sequences (references).
Reference sequences can be found with prefixes: "TTTTTT" for U6 (S pyogenes) barcode and "CGTGCAC" for H1 (S aureus) barcodes.

molecule: genomic DNA

growth_protocol: Cells were continuously cultured in RPMI 1640 supplemented with 10% FBS and penicilin/streptomycin.

extract_protocol:
gDNA was isolated with Qiagen Blood and Tissue kit according to the manufacturer's protocol.
PCR amplification was carried out with primers flanking sgRNA barcode pairs (Najm et al NBT 2017, Supplementary Note 1 for layout).

treatment_protocol: Cells carrying SpCas9 were transduced with lentivirus delivering SaCas9 and guide pair (300k or validation library). After 21 days, cells were harvested for gDNA extraction.

value: leukemia cell line


title: E12_LV_Tbx5_Ctrl_1


title: Reh-D

description:
Column name in the processed data file: Reh-D
300k_conditions.txt
300k_reference.txt
300k_counts.txt

source_name: Reh

value: Reh


title: THP-1-C

description:
Column name in the processed data file: THP-1-C
300k_conditions.txt
300k_reference.txt
300k_counts.txt


title: Reh-C

description:
Column name in the processed data file: Reh-C
300k_conditions.txt
300k_reference.txt
300k_counts.txt


title: 40k_library_screen

description: Column name in the processed data file: pDNA or cell line

data_processing:
Data were processed with PoolQ to split sample reads by Illumina barcode (conditions) and count sgRNA sequences (references).
Reference sequences require prefixes: "CACCG" within the first 30 bases for U6 (S pyogenes) driven sgRNAs and "CTTAAAC" for H1 (S aureus) driven sgRNAs.

extract_protocol:
gDNA was isolated with Qiagen Blood and Tissue kit according to the manufacturer's protocol.
PCR amplification was carried out with primers flanking sgRNA pair cassette (Najm et al NBT 2017, Supplementary Note 1 for primer sequences).

treatment_protocol: Cells carrying SpCas9 were transduced with lentivirus delivering SaCas9 and guide pair (40k library). After 21 days, cells were harvested for gDNA extraction.

value: leukemia cell lines


title: MTA1&2_A_MV4-11

description: Column name in the processed data file: MR76

data_processing:
Data were aligned with STAR aligner 43 to the hg38 genome assembly using the “Basic” two-pass mode. The option “--outFilterScoreMinOverLread“ was set to 0.2 to allow the mapping of shorter fragments
Raw gene counts were obtained using STAR aligner by adding the option “--quantMode GeneCounts” to the mapping script. NCBI RefSeq GTF file was used for gene counting purposes.
The merged length of all exons for each protein-coding gene was used to correct raw counts. TPM was calculated using the following formula in R: “tpm.mat <- t( t(x) * 1e6 / colSums(x) )” where x is the gene length-corrected expression matrix
Assembly: hg38
Supplementary files format and content: Tab-delimited text file including TPM valuies for each gene in all processed mRNA-Seq libraries

extract_protocol:
RNA was isolated with Qiagen RNeasy Plus Micro kit according to the manufacturer's protocol and ensuring RIN values were greater than 7.
Libraries were prepared first with Poly-A enrichment using magnetic oligo(dT)-beads (Invitrogen), then ligated to RNA adaptors for sequencing.

treatment_protocol: Cells were transduced with lentivirus delivering CRISPR-Cas9, guide, and fluorescent reporter. After 7 days, cells were flow analyzed, sorted, and collected for RNA extraction.

value: cell line

value: MTA1&2 KO


title: CHD4_B_MV4-11

description: Column name in the processed data file: MR79

value: CHD4 KO


title: MTA2_B_MV4-11

description: Column name in the processed data file: MR75

value: MTA2 KO


title: MTA2_A_MV4-11

description: Column name in the processed data file: MR74


title: MTA1_B_MV4-11

description: Column name in the processed data file: MR73

value: MTA1 KO


title: CHD4_A_MV4-11

description: Column name in the processed data file: MR78


title: MTA1_A_MV4-11

description: Column name in the processed data file: MR72


title: MTA1&2_B_MV4-11

description: Column name in the processed data file: MR77


title: Safe_B_MV4-11

description: Column name in the processed data file: MR71

value: Control


title: MTA1&2_B_Reh

description: Column name in the processed data file: MR67


title: validation_library_screen

description:
Column name in the processed data file: pDNA or cell line name
validation_conditions.txt
validation_reference.txt
validation_counts.txt


title: CHD4_B_Reh

description: Column name in the processed data file: MR69


title: E12_LV_Tbx5_Ctrl_3


title: MTA2_B_Reh

description: Column name in the processed data file: MR65


title: MTA1&2_A_Reh

description: Column name in the processed data file: MR66


title: MTA1_B_Reh

description: Column name in the processed data file: MR63


title: MTA1_A_Reh

description: Column name in the processed data file: MR62


title: CHD4_A_THP-1

description: Column name in the processed data file: MR58


title: CHD4_B_THP-1

description: Column name in the processed data file: MR59


title: Safe_A_MV4-11

description: Column name in the processed data file: MR70


title: Safe_A_Reh

description: Column name in the processed data file: MR60


title: MTA2_A_Reh

description: Column name in the processed data file: MR64


title: MTA2_A_THP-1

description: Column name in the processed data file: MR54


title: MTA2_B_THP-1

description: Column name in the processed data file: MR55


title: MTA1_B_THP-1

description: Column name in the processed data file: MR53


title: Safe_B_THP-1

description: Column name in the processed data file: MR51


title: Safe_A_THP-1

description: Column name in the processed data file: MR50


title: SFV_mutAA in C57BL/6_Brain #1

description:
SFV4 strain (GenBank: KP699763.1)
small RNA

source_name: C57/BL6 mice

value: brain

value: Semliki Forest virus


title: MTA1_A_THP-1

description: Column name in the processed data file: MR52


title: MTA1&2_A_THP-1

description: Column name in the processed data file: MR56


title: SFV in MLF_IFNAR1 KO

value: lung


title: Safe_B_Reh

description: Column name in the processed data file: MR61


title: AM_26C_IAV

city: Marburg

first: Wilhelm

last: Bertrams

address: Hans-Meerwein-Str. 2

department: Institute for Lung Research

institute: Philipps University Marburg

description:
AM from lung tissue
21_AM_26C_IAV

data_processing:
de-multiplexed fastq files were tested with FastQC. Over-represented sequences and adapters were trimmed with the Qiagen CLC Workbench Software v. 10.0.1.
Reads were mapped with the Qiagen CLC Workbench Software v. 10.0.1., using the hg19 human reference genome using default mapping parameters
Mapped total reads were analysed with the R package DeSeq2 v. 1.22.2 in R version 3.5.1, and the contrasts IAV_A549_vs_ctrl_A549, IAV_AEC_vs_ctrl_AEC, IAV_AM_vs_ctrl_AM and IAV_HuLu_vs_ctrl_HuLu  were calculated
The DeSeq2 output, namely the contrasts mentioned, are provided as excel file processed_results.xlsx
Assembly: hg19
Supplementary files format and content: processed_results.xlsx, DeSeq2 contrasts IAV_A549_vs_ctrl_A549, IAV_AEC_vs_ctrl_AEC, IAV_AM_vs_ctrl_AM and IAV_HuLu_vs_ctrl_HuLu

source_name: lung tissue

growth_protocol: A549 cells (ATCC, CCL-185) were cultured in Ham’s F12 medium supplemented with 10% fetal calf serum (FCS) and 2 mM L-glutamine at 37 °C in a 5% CO2 atmosphere.  Normal peripheral human lung tissue was freshly obtained from patients suffering from bronchial carcinoma.   AMs were isolated by repeated perfusion of the human lung tissue with Hanks' balanced salt solution (HBSS). The resulting cells were seeded in six-well plates (1 × 10^6 cells per well) and cultured in Roswell Park Memorial Institute (RPMI) 1640 medium supplemented with 2% FCS and 2 mM L-glutamine under the conditions described above.  AECIIs were isolated as follows: human lung tissue was perfused to remove AMs as stated above and the remaining tissue was finely minced and digested with type I trypsin and DNase. The digested tissue was filtered and cells were incubated in hybridoma serum-free medium (SFM) to promote the differential adhesion of residual AMs. The cell fraction was then collected and layered on a Pancoll discontinuous gradient. Cells at the interfacial layer were collected, washed with HBSS and finally 1 × 10^6 cells per well were seeded in a 24-well plate and cultured in RPMI 1649 medium supplemented with 10% FCS and 2 mM L-glutamine

extract_protocol:
RNA was isolated by phenol-chloroform extraction with Trizol Reagent followed by DNaseI digestion.
RNA libraries were prepared for sequencing using standard Illumina protocols.

treatment_protocol: Infection with Influenza A (A/Panama/2007/1999) was done at a multiplicity of infection (MOI) of 1 for 24 h.

tag: infection status

value: IAV infected

value: AM from lung tissue


title: A549_V1_C

description:
cell line
18_A549_V1_C

source_name: cell line

value: uninfected

value: A549 cell line


title: A549_V3_C

description:
cell line
17_A549_V3_C


title: AM_124V-2_IAV

description:
AM from lung tissue
19_AM_124V-2_IAV


title: AM_124V-1_C

description:
AM from lung tissue
23_AM_124V-1_C


title: SFV_K124A/K128A in MLF #3


title: AM_17B001_IAV

description:
AM from lung tissue
22_AM_17B001_IAV


title: AM_17B001_C

description:
AM from lung tissue
24_AM_17B001_C


title: SFV_K124A/K128A in MLF #1


title: AM_26C_C

description:
AM from lung tissue
20_AM_26C_C


title: MTA1&2_B_THP-1

description: Column name in the processed data file: MR57


title: SFV_mutAA in MLF #1


title: SFV_mutAA in MLF #2


title: A549_V2_IAV

description:
cell line
15_A549_V2_IAV


title: SFV in C57BL/6_Brain #1


title: A549_V1_IAV

description:
cell line
16_A549_V1_IAV


title: A549_V2_C

description:
cell line
14_A549_V2_C


title: SFV_mutAA in C57BL/6_Brain #2


title: SFV in C57BL/6_Brain #2


title: SFV in MLF_AGO-IP #3


title: AEC_6C_C

description:
AEC from lung tissue
12_AEC_6C_C

value: AEC from lung tissue


title: SFV_K124A/K128A in MLF #2


title: SFV in MLF #3


title: AEC_6C_IAV

description:
AEC from lung tissue
11_AEC_6C_IAV


title: AEC_213M_C

description:
AEC from lung tissue
10_AEC_213M_C


title: SFV in MLF_AGO-IP #2


title: SFV in MLF #2


title: SFV_mutAA in MLF #3


title: AEC_118V_IAV

description:
AEC from lung tissue
8_AEC_118V_IAV


title: SFV in MLF_AGO-IP #1


title: SFV in MLF #1


title: K2

city: Suzhou

first: Qiao

last: Cheng

state: Jiangsu Province

address: Shizi Street

department: Institute of Blood and Marrow Transplantation (IBMT)

institute: The First Affiliated Hospital of Soochow University

description: 10x Genomics

data_processing:
bcl2fastq tools for base calling
CellRanger software for single cell identifying
PCA and UMAP/TSNE for dimensionality reduction
Seurat for finding clusters
Assembly: mm10
Supplementary files format and content: Gene expression text files with tab-delimited

source_name: spleen

extract_protocol:
The totla cDNA was extracted  for libraries construction by Chromium  GEM Single Cell 3ʹ Reagent Kits v3 according the standard protocol
Libraries were prepared for sequencing using standard Illumina protocols
Single Cell 3ʹ Gene Expression libraries comprise standard Illumina paired-end   constructs which begin with P5 and end with P7. 16  bp 10x Barcodes are encoded at the start of TruSeq Read 1, while 8 bp sample index sequences are incorporated as the sample index read.

tag: strain

value: Balb/c

value: spleen


title: HuLu_549D_C

description:
whole lung tissue
5_HuLu_549D_C

value: whole lung tissue


title: AEC_213M_IAV

description:
AEC from lung tissue
7_AEC_213M_IAV


title: K1

description: 10x Genomics


title: HuLu_549D_IAV

description:
whole lung tissue
6_HuLu_549D_IAV


title: AEC_118V_C

description:
AEC from lung tissue
9_AEC_118V_C


title: W1

description: 10x Genomics

value: wild type


title: HuLu_221M_C

description:
whole lung tissue
1_HuLu_221M_C


title: HuLu_861Z_IAV

description:
whole lung tissue
2_HuLu_861Z_IAV


title: WT-Dox

city: Koto-ku

first: Tomoyoshi

last: Nakadai

state: Tokyo

country: Japan

institute: Cancer Institute of JFCR

data_processing:
Trimming adaptor sequences by skewer; Command line: skewer -f sanger -t 12 -m pe -x adaptor_sequences_files.fa --quiet -o ${name} ${name}_R1.fastq ${name}_R2.fastq
Quality check by fastqc; Command line: fastqc -o fastqc -f fastq --nogroup ${name}-trimmed-pair1.fastq ${name}-trimmed-pair2.fastq
Pre-alignment of reads to rCRSd to remove mitochondrial sequences by bowtie2 and samtools; Command line:(1) bowtie2 -p 12 -x rCRSd -k 1 -D 20 -R 3 -N 1 -L 20 -i S,1,0.50 -X 2000 --no-mixed --no-discordant -1 ${name}-trimmed-pair1.fastq -2 ${name}-trimmed-pair2.fastq -S ${name}_pre1.sam, (2) samtools sort -@ 12 -m 2G -n ${name}_pre1.sam > ${name}_pre1.sort.bam, (3) samtools view -@ 12 -S -f4 ${name}_pre1.sort.bam > ${name}_pre1.unmapped.sam, (4) cat ${name}_pre1.unmapped.sam | grep -v ^@ | awk 'NR%2==1 {print "@"$1"\n"$10"\n+\n"$11}' > ${name}_pre1_r1.fastq, and then (5) cat ${name}_pre1.unmapped.sam | grep -v ^@ | awk 'NR%2==0 {print "@"$1"\n"$10"\n+\n"$11}' > ${name}_pre1_r2.fastq
Pre-alignment of reads to human repeats to remove those sequences by bowtie2 and samtools; Command line: (1) bowtie2 -p 12 -x human_repeats -k 1 -D 20 -R 3 -N 1 -L 20 -i S,1,0.50 -X 2000 --no-mixed --no-discordant -1 ${name}_pre1_r1.fastq -2 ${name}_pre1_r2.fastq -S ${name}_pre2.sam, (2) samtools sort -@ 12 -m 2G -n ${name}_pre2.sam > ${name}_pre2.sort.bam, (3) samtools view -@ 12 -S -f4 ${name}_pre2.sort.bam > ${name}_pre2.unmapped.sam, (4) cat ${name}_pre2.unmapped.sam | grep -v ^@ | awk 'NR%2==1 {print "@"$1"\n"$10"\n+\n"$11}' > ${name}_pre2_r1.fastq, and then (5) cat ${name}_pre2.unmapped.sam | grep -v ^@ | awk 'NR%2==0 {print "@"$1"\n"$10"\n+\n"$11}' > ${name}_pre2_r2.fastq
Alignment of reads to hg38 by bowtie2; Command line: bowtie2 -p 12 -x hg38 --very-sensitive -X 2000 --no-mixed --no-discordant -1 ${name}_pre2_r1.fastq -2 ${name}_pre2_r2.fastq -S ${name}_align.sam
Removing low quality reads by samtools; Command line: (1) samtools view -bS -@ 12 ${name}_align.sam > ${name}_align.bam (2) samtools sort -@ 12 ${name}_align.bam > ${name}_align.sort.temp.bam, and then (3) samtools view -f 2 -q 10 -b -@ 12 -m 2G ${name}_align.sort.temp.bam > ${name}_align.sort.bam
Removing PCR duplicates by Picard; Command line: java -jar picard.jar MarkDuplicates I=${name}_align.sort.bam O=${name}.rmdup.bam M=metrics.file VALIDATION_STRINGENCY=LENIENT REMOVE_DUPLICATES=true
Peak calling by MACS2; Command line: macs2 callpeak -t ${name}_align.sort.bam -n ${name} -f BAM -g hs --shift -75 --extsize 150 --nomodel --call-summits --nolambda --keep-dup all -q 0.01 --outdir peak
Genome_build: hg38
Supplementary_files_format_and_content: narrowPeak

growth_protocol: MDA-MB-453 cells inducibly expressing wild-type, Y537S, or K303R ER alpha (1 x 10^6 cells) were plated on a 100 mm tissue culture plate with McCoy's 5A medium supplemented with 10% FBS and 1% Penicillin/Streptomycin, cultured for 2 days, added with/without 2 ug/mi doxycycline and/or 1nM E2, further cultured for 2 days, and then harvested for ATAC-seq.

extract_protocol:
5 x 10^4 cells are processed for nuclear pellet preparation with a hypotonic buffer with digitonin.
Libraries were prepared from the nuclear pellets by using Tagment DNA Enzyme kit (Illumina). Prepared DNA fragments were cleaned with DNA Clean and concentrator-5 kit (Zymo Research) and added with adaptor and index sequences by PCR with NEBNext high-fidelity 2x PCR master mix (NEB), and then purified with DNA Clean and concentrator-5 kit (Zymo Research) again. The concentration of the library was quantified using Fluorometer (Quantus) and KAPA library quantification kit (Roche), and the quality was checked by TapeStation D5000 (Agilent). The pooled libraries with unique index sequences were sequenced using MiSeq or NextSeq550 (Illumina) with pair-end 75 bp reads following the manufacturer's protocols.

value: treated with doxycycline for 2 days


title: W2

description: 10x Genomics


title: HuLu_861Z_C

description:
whole lung tissue
4_HuLu_861Z_C


title: HuLu_221M_IAV

description:
whole lung tissue
3_HuLu_221M_IAV


title: WT-DoxE2

value: treated with doxycycline/E2 for 2 days


title: A549_V3_IAV

description:
cell line
13_A549_V3_IAV


title: FRA102220_005

city: Chicago

first: Leonid

last: Feferman

state: Illinois

address: 1737 West Polk St

department: Research Resources Center

institute: University of Illinois at Chicago

description:
single-cell (10Xgenomics)
Cells were seeded in low attachment plates at single-cell density and were grown in mammoshpere conditions for 7 days.

data_processing:
Raw data was processed and aligned by the UIC Research Informatics. cellRanger (v3.0.0) was run on the raw data to align Ensembl genome GRCh38 for 10X Genomics data Unique UMI counts were summed for each gene and each unique cell barcode.   The output was analyzed using the standard workflow from Seurat package (Version 4.0.6) in the Rstudio (v.4.0.3). Cell duplicates, cells with low gene counts (<2000 genes), and cells with high mitochondrial gene expression (>15% of total mapped reads) were excluded from analysis.
Assembly: hg38 Ensembl gene sequences (exonic only)
Supplementary files format and content: barcodes.tsv.gz, features.tsv.gz, matrix.mtx.gz

source_name: Breast

growth_protocol: Before mammosphere assay, cells were routinely cultured in RPMI 1640 containing  10% fetal bovine serum, 1% penicillin–  streptomycin, 1% nonessential amino acids, 2 mmol/L L-glutamine, and 0.5 µg/ml puromycin.

extract_protocol:
For scRNA-seq, MS were collected, washed twise with 1x PBS, trypsinized and resuspended in x1 PBS with 0.04% BSA. Cell suspension was loaded on a Chromium Single Cell 3′ Chip (10X Genomics).
Single-cell libraries were prepared according to the manufacturer’s protocol. 10X libraries were pooled and were sequenced on an S4 lane with 28x150nt reads and produced over 5.4 billion reads.

value: T47D cell line

value: Breast

value: Cultured in mammosphere conditions


title: huTNFtg;TgColVI-miR-221/222, rep3

city: Vari

first: George

last: Kollias

state: Attica

address: 34 Fleming Street

department: Institute for Bioinnovation/ Department of Physiology, School of Medicine

country: Greece

institute: BSRC "Alexander Fleming"/ National and Kapodistrian University of Athens

data_processing:
Quality of the FASTQ files (obtained after Ion Proton sequencing) was assessed by FastQC v0.11.8.
Alignment of sequencing reads to the mouse reference genome was performed by the software HISAT2 (version 2.1.0 and parameters: --score-min L,0,-0.36 -k 2 --max-seeds 20 --rna-strandness F -p 6) with the mm10 version of the reference genome.
The bam files were summarized to read counts tables using FeatureCounts (version 1.6.0, and parameters: -t 'three_prime_utr' -g 'gene_name' --largestOverlap --readExtension3 500 --readExtension5 500 -s 1) and the gene annotation file mus_musculus.grcm38.92.gtf from Ensembl database.
The resulting gene counts tables were subjected to differential expression analysis (DEA) utilizing the R package DESeq2 (v1.34.0). Differentially expressed genes were identified after setting the following thresholds: pvalue < 0.05 and |log2FC| > 1.
Two samples (Wt3 and Tg197-C2) were excluded from analysis, because they did not match the quality control criteria.
Assembly: mm10 (from Illumina iGenomes)
Supplementary files format and content: Raw gene count table for each sample in .txt format.

source_name: ankle joints

extract_protocol:
Total RNA was extracted using RNAeasy Mini kit (QIAGEN), according to the manufacturer's instructions.
The quantity and quality of RNA samples from sorted cells were analyzed using the bioanalyzer form Agilent in combination with the Agilent RNA 6000 Nano. RNA samples with RNA Integrity Number (RIN) > 7 were further used for library preparation using the 3’ mRNA-Seq Library Prep Kit Protocol for Ion Torrent (QuantSeq-LEXOGEN™) according to manufacturer’s instructions. The quantity and quality of libraries were assessed using the DNA High Sensitivity Kit in the bioanalyzer, according to the manufacturer’s instructions (Agilent Technologies). Libraries were subsequently pooled and templated using the Ion PI IC200 Chef Kit (ThermoFisher Scientific) on an Ion Proton Chef instrument or Ion OneTouch System. Sequencing was performed using the Ion PI™ Sequencing 200 V3 Kit and Ion Proton PI™ V2 chips (ThermoFisher Scientific) on an Ion Proton™ System, according to the manufacturer's instructions.

value: ankle joints

value: Synovial fibroblasts


title: TgColVI-miR-221/222, rep3


title: FRA072420_001

description:
single-cell (10XGenomics)
Cells were seeded in low attachment plates at single-cell density and were grown in mammoshpere conditions for 7 days.

value: MCF-7 cell line


title: TgColVI-miR-221/222, rep2


title: U2OS_R2_Late S Fraction

city: Minneapolis

first: Juan Carlos

last: Rivera-Mulia

state: MN

address: 420 Washington Ave SE

department: Department of Biochemistry, Molecular Biology and Biophysics

institute: University of Minnesota Medical School

data_processing:
Library strategy: Repli-seq
Reads were mapped to corresponding genome (hg38 or mm10) using BWA.
Coverage data are calculated from mapped reads using samtools and bedtools
Base 2 log of early to late coverage ratio was calculated in 20kb windows
Log 2 ratio data were quantile normalized using R package preprocessCore
Data were loess smoothed and binned into 10kb windows
Genome_build: hg38 and mm10
Supplementary_files_format_and_content: processed data are 10kb windows in hg38 or mm10 genomes.

source_name: Osteosarcoma cells

growth_protocol: H9 cells were cultured in mTeSR Plus (StemCell Technologies Cat. No. 05825) on Geltrex (Life Technologies Cat. No. A1413302) coated dishes according to WiCell instructions. U2OS cells were grown in DMEM media (Corning Cat.No. 15013CM) supplemented with 10% FBS (Corning Cat.No.35011CV), 1% Glutamine (Gibco Cat.No. 25030081), 1% Penicillin-Streptomycin (Gibco Cat.No. 15070-063). MEFs were cultured in 1:1 DMEM media (Corning Cat.No. 10-017-CV) and F10 media with Glutamax (Gibco Cat.No. 41550021), supplemented with 10% FBS (Sigma Cat.No. F2442), 1X NEAA (Gibco Cat.No. 11140050) and 1% Penicillin-Streptomycin (Corning Cat.No. 30002Cl).

extract_protocol:
Genome-wide replication timing profiles were constructed as previously described (Hiratani et al. 2008; Ryba et al. 2011; Marchal et al., 2017). Briefly, cells were pulse labeled with BrdU and separated into early and late S-phase fractions by flow cytometry and processed by NGS.
Sequencing libraries of BrdU-substituted DNA from early and late fractions were prepared by NEBNext Ultra DNA Library Prep Kit for Illumina (E7370) or by KAPA HyperPlus library preparation kit (Roche Cat. No.  07958978001).

tag: s-phase fraction

value: Late S fraction

tag: replicate


title: U2OS_R2_Early S Fraction

value: Early S fraction


title: U2OS_R1_Late S Fraction


title: TgColVI-miR-221/222, rep1


title: huTNFtg, rep3

value: huTNFtg


title: huTNFtg, rep1


title: H9_R1_Early S Fraction

source_name: Human ESC


title: huTNFtg, rep2


title: CYTO_10

city: Indianapolis

first: Carmella

last: Evans-Molina

state: Indiana

address: 1210 Waterway Blvd, 4108G, Indianapolis, IN

department: Pediatrics

institute: Indiana University School of Medicine and the Center for Diabetes and Metabolic Diseases

description:
Evans-Molina_Splicing_SE.MATS.txt
Evans-Molina_Splicing_A5SS.MATS.txt
Evans-Molina_Splicing_A3SS.MATS.txt
Evans-Molina_Splicing_MXE.MATS.txt
Evans-Molina_Splicing_RI.MATS.txt

data_processing:
Sequence reads were aligned to the human genome (GENCODE GRCh37) using the STAR RNA-seq aligner.Read distributions across the genome were assessed using bamutils (from ngsutils)
Alternative splicing events were analyzed using replicated multivariate analysis of transcript splicing (rMATs),which computes a percentage splicing index (PSI) and false discovery rate (FDR) for five different splicing events, including skipped exons, mutually exclusive exons, retained introns, and 5’ and 3’ alternative splice sites.
Genome_build: GRCh37
Supplementary_files_format_and_content: tab-delimited text files include splicing output from rMATs.

source_name: Human islets

extract_protocol:
SeraMir RNA isolation kit (SBI Biosciences)
RNA quantification and quality were assessed by measuring the 260/280 ratio using a Nanophotometer (Implen, Munich, Germany). RNA integrity was evaluated using the Bioanalyzer 2100 (Agilent Technologies, Santa Clara, USA).  The average (S.D.) RIN value for islet samples utilized in this study was 9.8  0.28. Next, 500 nanograms of RNA were used to prepare a single-indexed strand-specific cDNA library using a TruSeq Stranded mRNA Library Prep Kit (Illumina). The resulting libraries were assessed for quantity and size distribution using a Qubit and Agilent 2100 Bioanalyzer; 1.5 pM pooled libraries were sequenced with 2×75bp paired-end configuration on an Illumina NextSeq500 using a NextSeq 500/550 High Output Kit with an average 52.6M reads. A Phred quality score (Q score) was used to measure the quality of sequencing, and >90% of the sequencing reads reached Q30 (99.9% base call accuracy).

treatment_protocol: Upon receipt, islets were cultured overnight in standard Prodo medium (Prodo Labs, CA) and then treated with or without 50 U/ml IL-1β and 1000 U/ml IFN-γ for 24 hours.

value: Human islets from cadaveric organ donors

value: Treated with IL-1β and IFN-γ for 24 hours


title: H9_R1_Late S Fraction


title: wt, rep2

value: wt


title: CYTO_8


title: CYTO_6


title: SFRP2 overexpressed PC3 cells isolated from COL1 Coated Surface-Biol Rep2

city: München

first: Elif

last: Akova Ölken

state: Baveria

address: Fraunhofer Str. 20

department: Department of Orthopaedics and Trauma Surgery

institute: Ludwig-Maximilians-University

data_processing:
FASTA file demultiplexed on Galaxy
STAR used for alignment
Read per gene count  files merged as raw data for analysis
Normalised raw file created on R software
Assembly: GRCh38.101

source_name: prostate cancer

growth_protocol: Dulbecco´s Modified Eagle Medium + GlutaMax (Thermo Fisher, USA) with 10% fetal bovine serum (FBS, Sigma Aldrich, USA) and 1% penicillin/streptomycin (Sigma Aldrich, USA) at 37°C with 5% CO2.

extract_protocol:
To identify SFRP2‑induced transcriptional changes in PC3 cells, 100,000 cells were seeded T25 flasks and RNA was isolated after 5 days and RNA integrity was validated with Bioanalyzer (Agilent, USA)
cDNA sequencing libraries were prepared with a SENSE mRNA-Seq Library Prep Kit V2 (Lexogen, Austria).

treatment_protocol: Cells seeded on Nunc and COL1 coated surface

value: prostate cancer

value: SFRP2 OE

value: COL1 coated surface


title: SFRP2 overexpressed PC3 cells isolated from COL1 Coated Surface-Biol Rep3


title: huTNFtg;TgColVI-miR-221/222, rep1


title: SFRP2 overexpressed PC3 cells isolated from COL1 Coated Surface-Biol Rep1


title: PC3 Cells isolated from COL1 coated Surface-Biol Rep2


title: CYTO_5


title: CYTO_4


title: PC3 Cells isolated from COL1 coated Surface-Biol Rep1


title: CYTO_3


title: SFRP2 overexpressed PC3 cells isolated from Nunc Surface-Biol Rep3

value: Nunc


title: CYTO_2


title: CYTO_1


title: U2OS_R1_Early S Fraction


title: CONT_10

value: Untreated control


title: SFRP2 overexpressed PC3 cells isolated from Nunc Surface-Biol Rep1


title: wt, rep1


title: CYTO_9


title: CONT_9


title: PC3 Cells isolated from Nunc Surface-Biol Rep2


title: PC3 Cells isolated from Nunc Surface-Biol Rep3


title: CONT_8


title: CONT_7


title: PC3 Cells isolated from COL1 coated Surface-Biol Rep3


title: CONT_6


title: CYTO_7


title: CONT_5


title: CONT_4


title: PC3 Cells isolated from Nunc Surface-Biol Rep1


title: CONT_3


title: CONT_2


title: Case 120: type B3 thymoma

type: genomic

city: Heidelberg

first: Daniel

last: Schrimpf

address: Im Neuenheimer Feld 224

department: Neuropathology

institute: University Hospital Heidelberg, Intitute of Pathology

description: type B3 thymoma

hyb_protocol: Illumina protocol

scan_protocol: Illumina iScan System according to the manufacturer's protocol

data_processing: Bioconductor (Version 3.3) package minifi (Version 1.18) using function 'preprocessIllumina'  Average beta

label: Cy3 and Cy5

source_name: type B3 thymoma

label_protocol: Illumina protocol

extract_protocol: Illumina protocol

treatment_protocol: An automated extraction was done with a Maxwell system (Promega, Fitchburg, WI, USA) and the Maxwell 16 FFPE Plus LEV DNA Purification Kit according to the manufacturer’s instructions.

tag: material

value: FFPE


title: Case 122: type B3 thymoma

description: type B3 thymoma


title: Case 117: type B3 thymoma

description: type B3 thymoma


title: SFRP2 overexpressed PC3 cells isolated from Nunc Surface-Biol Rep2


title: Case 121: type B3 thymoma

description: type B3 thymoma


title: Case 118: type B3 thymoma

description: type B3 thymoma


title: Case 119: type B3 thymoma

description: type B3 thymoma


title: Case 116: type B3 thymoma

description: type B3 thymoma


title: CONT_1


title: Case 115: type B3 thymoma

description: type B3 thymoma


title: Case 114: type B3 thymoma

description: type B3 thymoma


title: KRAB Control on Day1

city: Ann Arbor

first: Dogukan

last: Mizrak

state: MI

address: NCRC Building 26, 263S

department: Cardiac Surgery

institute: University of Michigan

data_processing:
12-nt cell barcodes (CBs) and 8-nt unique molecular identifiers (UMIs) were extracted from read 1.
Read 2 were trimmed to remove poly(A) tails.
Trimmed reads were aligned to the human genome and transcriptome annotation (GRCh38) using STAR v.2.5 with parameters --sjdbOverhang 65 --twopassMode Basic --outSAMtype BAM Unsorted. Only reads with unique, strand-specific alignments to exons were kept for further analysis.
Genome_build: GRCh38
Supplementary_files_format_and_content: KRAB_Control_Day1_matrix.txt, KRAB_Control_Day3_matrix.txt and rs117430032i_Day3_matrix.txt contain the gene expression profiles of each cell. KRAB_Control_Day1_barcodes.txt, KRAB_Control_Day3_barcodes.txt, and rs117430032i_Day3_barcodes.txt contain the ordered cell barcodes for each cell in the gene expression matrix.

source_name: EndoMT

extract_protocol:
mRNA molecules were captured using bead-bound oligo dT(30).
Captured mRNA molecules were reverse transcribed with template switching to obtain cDNA. cDNA was amplified by PCR reactions (12 cycles). Aliquots of amplified cDNA from each sample were pooled, and purified with AMPure XP beads. The purified PCR products were used as input for the Nextera XT reactions to generate the sequencing libraries.

tag: gender

value: Male

tag: number of cells


title: Case 113: type B3 thymoma

description: type B3 thymoma


title: K303R-DoxE2 [ChIP-Seq]

data_processing:
Quality filtering by FASTX-toolkit; Command line: fastq_quality_filter -q 25 -p 90 -i ${name}_R1.fastq -o ${name}.qc.fastq
Alignment of reads to hg38 by bowtie2; Command line: bowtie2 -p 12 -x hg38 -U ${name}.qc.fastq -S ${name}.sam
Removing multi-mapped reads; Command line: (1) grep -v "XS" ${name}.sam > ${name}.uq.sam, (2) samtools view -Shb ${name}.uq.sam > ${name}.uq.bam, and then (3) samtools sort -@ 12 -m 2G ${name}.uq.bam > ${name}.uq.sort.bam
Removing duplicated reads by Picard; Command line: java -jar picard.jar MarkDuplicates INPUT=${name}.uq.sort.bam OUTPUT=${name}.drm.bam METRICS_FILE=${name}.out.metrics AS=true REMOVE_DUPLICATES=true VALIDATION_STRINGENCY=LENIENT
Removing reads included in the black list by Bedtools; Command line: intersectBed -abam ${name}.drm.bam -b hg38.blacklist.bed –v  >${name}.drm.blalst.bam
Peak calling by MACS2; Command line; macs2 callpeak -t ${name}.drm.blalst.bam -n ${name}.drm.blalst -g hs --keep-dup auto
Genome_build: hg38
Supplementary_files_format_and_content: narrowPeak

growth_protocol: MDA-MB-453 cells inducibly expressing wild-type, Y537S, or K303R ER alpha (2.6 x 10^6 cells) were plated on a 100 mm tissue culture plate with McCoy's 5A medium supplemented with 10% FBS and 1% Penicillin/Streptomycin, cultured for 2 days, added with/without 2 ug/mi doxycycline and/or 1nM E2, further cultured for 2 days, and then used for ChIP-seq.

extract_protocol:
The cells on two 100 mm plates were fixed and nuclear pellets were sonicated to prepare soluble chromatin lysate. ER alpha-chromatin complex was immunoprecipitated by anti-ER alpha antibody from the lysate and precipitated DNA was recovered by reverse crosslinking.
The precipitated DNA was purified with AMPure XP beads (Agencourt) and ChIP-seq library was prepared from 2 ng of the purified DNA using SMARTer ThruPLEX DNA-seq Kit (TAKARA). The library was then purified using AMPure XP beads (Agencourt) again. The concentration of the library was quantified using Fluorometer (Quantus) and KAPA library quantification kit (Roche), and the quality was checked by TapeStation D5000 (Agilent). The pooled libraries with unique index sequences were sequenced using NextSeq550 (Illumina) with pair-end 75 bp reads following the manufacturer's protocols.

tag: cell line

value: treated with doxycycline/E2 for 2 days

tag: chip antibody

value: Anti-ERa (D8H8), CST Japan, #8644


title: Case 111: type B3 thymoma

description: type B3 thymoma


title: Case 110: type B2 thymoma

description: type B2 thymoma

source_name: type B2 thymoma


title: KRAB Control on Day3


title: rs117430032i on Day3

value: dcas9-KRAB-MeCP2 + rs117430032 gRNA


title: Case 109: type B2 thymoma

description: type B2 thymoma


title: Case 108: type B2 thymoma

description: type B2 thymoma


title: K303R-Dox [ChIP-Seq]

tag: treatment

value: treated with doxycycline for 2 days


title: WT-DoxE2 [ChIP-Seq]


title: Y537S-Dox [ChIP-Seq]


title: WT-Dox [ChIP-Seq]


title: Y537S-DoxE2 [ChIP-Seq]


title: K303R-DoxE2


title: Case 106: type B2 thymoma

description: type B2 thymoma


title: Case 105: type B2 thymoma

description: type B2 thymoma


title: Case 103: type B2 thymoma

description: type B2 thymoma


title: Case 104: type B2 thymoma

description: type B2 thymoma


title: Y537S-DoxE2


title: Case 102: type B2 thymoma

description: type B2 thymoma


title: K303R-E2

value: treated with E2 for 2 days


title: Y537S-Dox


title: K303R-NT

value: untreated


title: Case 101: type B2 thymoma

description: type B2 thymoma


title: K303R-Dox


title: Y537S-E2


title: Case 100: type B2 thymoma

description: type B2 thymoma


title: Case 112: type B3 thymoma

description: type B3 thymoma


title: Case 99: type B2 thymoma

description: type B2 thymoma


title: Y537S-NT


title: WT-E2


title: WT-NT


title: Case 98: type B2 thymoma

description: type B2 thymoma


title: Case 97: type B2 thymoma

description: type B2 thymoma


title: Case 107: type B2 thymoma

description: type B2 thymoma


title: RE8_T

city: Beijing

first: Jiabo

last: Di

address: Fucheng Road 52

institute: Peking University Cancer Hospital

data_processing:
Paired-end clean reads were aligned to the human reference genome GRCh38 (hg38) using Hisat2 v2.0.5.
Counts v1.5.0-p3 was used to count the read numbers mapped to each gene.
Fragments per kilobase of transcript per million mapped reads (FPKM) of each gene were calculated based on the length of the gene and read count mapped it.
Genome_build: GRCh38
Supplementary_files_format_and_content: xls file includes FPKM values for each sample

source_name: colorectum

extract_protocol:
Trizol
Sequencing libraries were generated using NEBNext® UltraTM RNA Library Prep Kit for Illumina

tag: tissue

value: tumor


title: Case 96: type B2 thymoma

description: type B2 thymoma


title: Case 95: type B2 thymoma

description: type B2 thymoma


title: RE7_T


title: Case 93: type B2 thymoma

description: type B2 thymoma


title: RE7_N

value: adjacent normal


title: Case 94: type B2 thymoma

description: type B2 thymoma


title: RE6_T


title: Case 92: type B2 thymoma

description: type B2 thymoma


title: Case 91: type B2 thymoma

description: type B2 thymoma


title: RE6_N


title: RE5_T


title: RE5_N


title: Case 90: type B2 thymoma

description: type B2 thymoma


title: Case 87: type B2 thymoma

description: type B2 thymoma


title: Case 89: type B2 thymoma

description: type B2 thymoma


title: RE4_T


title: Case 88: type B2 thymoma

description: type B2 thymoma


title: RE4_N


title: Case 85: type B2 thymoma

description: type B2 thymoma


title: RE3_T


title: Case 84: type B2 thymoma

description: type B2 thymoma


title: Case 86: type B2 thymoma

description: type B2 thymoma


title: RE2_N


title: RE2_T


title: RE3_N


title: Case 82: type B2 thymoma

description: type B2 thymoma


title: RE1_T


title: RE8_N


title: Case 80: type B2 thymoma

description: type B2 thymoma


title: LC6_N


title: RE1_N


title: Case 81: type B2 thymoma

description: type B2 thymoma


title: Case 79: type B2 thymoma

description: type B2 thymoma


title: LC6_T


title: LC5_T


title: Case 78: type B2 thymoma

description: type B2 thymoma


title: LC5_N


title: LC4_T


title: LC4_N


title: Case 76: type B1 thymoma

description: type B1 thymoma

source_name: type B1 thymoma


title: Case 75: type B1 thymoma

description: type B1 thymoma


title: LC3_T


title: Case 74: type B1 thymoma

description: type B1 thymoma


title: Case 73: type B1 thymoma

description: type B1 thymoma


title: Case 72: type B1 thymoma

description: type B1 thymoma


title: LC2_T


title: Case 83: type B2 thymoma

description: type B2 thymoma


title: LC2_N


title: LC1_T


title: Case 69: type B1 thymoma

description: type B1 thymoma


title: LC1_N


title: Case 70: type B1 thymoma

description: type B1 thymoma


title: Case 71: type B1 thymoma

description: type B1 thymoma


title: RC6_T


title: RC6_N


title: Case 68: type AB thymoma

description: type AB thymoma

source_name: type AB thymoma


title: Case 66: type AB thymoma

description: type AB thymoma


title: Case 67: type AB thymoma

description: type AB thymoma


title: RC5_N


title: Case 65: type AB thymoma

description: type AB thymoma


title: RC4_T


title: Case 77: type B1 thymoma

description: type B1 thymoma


title: RC4_N


title: LC3_N


title: Case 63: type AB thymoma

description: type AB thymoma


title: RC3_N


title: RC2_T


title: Case 62: type AB thymoma

description: type AB thymoma


title: RC2_N


title: Case 60: type AB thymoma

description: type AB thymoma


title: Case 61: type AB thymoma

description: type AB thymoma


title: Case 59: type AB thymoma

description: type AB thymoma


title: RC1_T


title: Case 58: type AB thymoma

description: type AB thymoma


title: RC1_N


title: Case 56: type AB thymoma

description: type AB thymoma


title: Case 55: type AB thymoma

description: type AB thymoma


title: RC5_T


title: Case 54: type AB thymoma

description: type AB thymoma


title: Case 52: type AB thymoma

description: type AB thymoma


title: Case 64: type AB thymoma

description: type AB thymoma


title: 21-2hr-Eto_rep2_R1.fastq.gz

city: BOSTON

first: ELIZABETH

last: POLLINA

state: MA

address: 220 LONGWOOD AVENUE, GOLDENSON 405

department: Neurobiology

institute: HARVARD MEDICAL SCHOOL

description: ENDseq in cultured mouse cortical neurons with etoposide treatment at the indicated stimulation timepoint

data_processing:
Quality read trimming was performed using trimmomatic/0.36 using the following command: java -jar $TRIMMOMATIC/trimmomatic-0.36.jar SE -threads 1 -phred33 [FASTQ_FILE] ILLUMINACLIP:/n/app/trimmomatic/0.36/bin/adapters/TruSeq3-SE.fa:2:30:10 LEADING:5 TRAILING:5 SLIDINGWINDOW:4:20 MINLEN:45
Single-end reads were mapped to the mouse genome using subread with the following command: subread-align -i /[PATH TO GENOME REFERENCE]/ -t 0 -r |FASTQ_FILE}_trimmed.fq -o [FASTQ_FILE].sam --SAMoutput
Assembly: mm10

source_name: Cultured cortical neurons

extract_protocol:
To dissociate cultured mouse cortical neurons for END-seq, Papain (Worthington Biochemical Cat: LK003178) was dissolved in TrypLE Express Enzyme solution at 37 °C for 20 minutes prior to sample collection. Culture media was gently aspirated and the cells were gently washed once with PBS at 37 °C. 500uL of papain solution was added to each well of neurons in 6 well plates and incubated at 37 °C for 1 min. Papain solution was removed via pipette and 500uL trituration solution (culture medium supplemented with freshly dissolved DNase) was added. Cells were gently triturated 5-10 times with wide bore pipette tip, transferred to conical tubes, and pelleted. Cells were gently resuspended in ice cold PBS containing 0.1% BSA and 0.5 mM EDTA for counting. Cells were stored on ice while an aliquot was quickly counted via hemocytometer. 1.5 million neurons for each experimental condition were added to a new conical tube and pelleted. Cells were then embedded in agarose plugs using Bio-Rad CHEF Mammalian Genomic DNA Plug Kit (Bio-Rad #1703591) and processed for END-seq as previously described19,90. Experimental groups were processed back-to-back, collecting cells in one treatment group and embedding them in agarose plugs before collecting cells in the next treatment group, in order to reduce over-digestion of the samples and to minimize time between cell dissociation and embedding in agarose. We observed better signal-to-noise in cells treated with etoposide (50uM). Etoposide was added to the cells 6 hours prior to collection, and was included at 50uM in all solutions applied to the cells until embedding in agarose.
END-seq

tag: cell/tissue type

value: Cultured cortical neurons

tag: mouse strain

tag: timepoint

value: 2hr post KCl depolarization


title: 7-2hr-Eto_rep1_R1.fastq.gz


title: 18-0hr-Eto_rep2_R1.fastq.gz

value: Unstim (0hr)


title: 4-0hr-Eto_rep1_R1.fastq.gz


title: Case 50: type AB thymoma

description: type AB thymoma


title: Case 51: type AB thymoma

description: type AB thymoma


title: sBLISS_RNAseq_10hr-Rep9

description: RNA-seq at indiciated stimulation timepoint from mouse hippocampus, paried with sBLISS-seq data from the same sample

data_processing:
Quality read trimming was performed using trimmomatic/0.36 using the following command: java -jar $TRIMMOMATIC/trimmomatic-0.36.jar SE -threads 1 -phred33 [FASTQ_FILE] ILLUMINACLIP:/n/app/trimmomatic/0.36/bin/adapters/TruSeq3-SE.fa:2:30:10 LEADING:5 TRAILING:5 SLIDINGWINDOW:4:20 MINLEN:45
For primary neuron culture RNA-seq, single-end reads were mapped to the mouse genome using subread with the following command: subread-align -i /[PATH TO GENOME REFERENCE]/ -t 0 -r |FASTQ_FILE}_trimmed.fq -o [FASTQ_FILE].sam --SAMoutput
For hippocampal tissue RNA-seq paired with sBLISS-seq, paired-end reads were mapped to the mouse genome using subread with the following command: subread-align -i /[PATH TO GENOME REFERENCE]/ -t 0 -r [FASTQ_FILE]_R1_paired_trimmed.fq -R [FASTQ_FILE]_R2_paired_trimmed.fq -o [FASTQ_FILE].bam
The featureCounts package was used to count mapped reads in RNA-seq data using a custom filtered annotation file: gencode.v17.annotation.gtf filtered for feature_type=”gene”, gene_type="protein_coding" and gene_status="KNOWN") to obtain read counts along genes for each sample.
Assembly: mm10
Supplementary files format and content: Counts matrices are provided from featureCounts. Rows are genes and columns represent each indicated sample and bioreplicate.

source_name: Mouse hippocampus

extract_protocol:
Hippocampal tissue was collected in Trizol reagent, and RNA was extracted using the RNeasy Kit (Qiagen) according to the manufacturer’s instructions.
Total RNA (1000 ng) was used to generate libraries following rRNA depletion (NEBNext, E6310X) according to the manufacturer’s instructions (NEBNext, E7420). 40 bp paired-end reads were generated on the Illumina NextSeq 500.
Bulk total RNA-seq

value: Mouse hippocampus

value: 10hr post KA injection


title: Case 48: type AB thymoma

description: type AB thymoma


title: Case 49: type AB thymoma

description: type AB thymoma


title: Case 47: type AB thymoma

description: type AB thymoma


title: RC3_T


title: Case 46: type AB thymoma

description: type AB thymoma


title: Case 45: type AB thymoma

description: type AB thymoma


title: Case 44: type A thymoma

description: type A thymoma

source_name: type A thymoma


title: Case 43: type A thymoma

description: type A thymoma


title: Case 40: type A thymoma

description: type A thymoma


title: Case 39: type A thymoma

description: type A thymoma


title: Case 57: type AB thymoma

description: type AB thymoma


title: Case 41: type A thymoma

description: type A thymoma


title: Case 37: type A thymoma

description: type A thymoma


title: Case 38: type A thymoma

description: type A thymoma


title: Case 36: type A thymoma

description: type A thymoma


title: Case 35: normal thymus

description: normal thymus

source_name: normal thymus


title: Case 34: normal thymus

description: normal thymus


title: Case 53: type AB thymoma

description: type AB thymoma


title: Case 33: normal thymus

description: normal thymus


title: Case 32: normal thymus

description: normal thymus


title: Case 31: normal thymus

description: normal thymus


title: Case 30: normal thymus

description: normal thymus


title: Case 29: normal thymus

description: normal thymus


title: sBLISS_RNAseq_0hr-Rep2

value: 0hr (PBS injected)


title: Case 27: normal thymus

description: normal thymus


title: Case 28: normal thymus

description: normal thymus


title: 6hr_p400_480_Rep3

description: RNA-seq at indiciated stimulation timepoint with indicated shRNA expressed

extract_protocol:
Neurons infected with either control shRNA virus or viruses targeting Npas4, EP400, or Kat5/Tip60 were washed twice with PBS to remove dead cells and scraped immediately into Trizol. Tissue was collected in Trizol reagent, and RNA was extracted using the RNAeasy Kit (Qiagen) according to the manufacturer’s instructions.
Total RNA (1000 ng) was used to generate libraries following rRNA depletion (NEBNext, E6310X) according to the manufacturer’s instructions (NEBNext, E7420). Eighty-five bp reads were generated on the Illumina Nextseq 500
Bulk total RNA-seq

value: 6hr post KCl depolarization


title: Case 26: micronodular thymoma

description: micronodular thymoma

source_name: micronodular thymoma

value: FFPE


title: Case 25: micronodular thymoma

description: micronodular thymoma


title: Case 42: type A thymoma

description: type A thymoma


title: 6hr_Tip60_300_Rep3


title: 6hr_p400_315_Rep3


title: Case 24: micronodular thymoma

description: micronodular thymoma


title: Case 23: micronodular thymoma

description: micronodular thymoma


title: 6hr_Tip60_299_Rep3


title: Case 22: micronodular thymoma

description: micronodular thymoma


title: 6hr_N4shRNA_Rep3


title: 2hr_p400_480_Rep3


title: Case 20: micronodular thymoma

description: micronodular thymoma


title: Case 21: micronodular thymoma

description: micronodular thymoma


title: 6hr_Ctrl_Rep3


title: 2hr_Tip60_300_Rep3


title: 2hr_Tip60_299_Rep3


title: Case 18: micronodular thymoma

description: micronodular thymoma


title: Case 19: micronodular thymoma

description: micronodular thymoma


title: 2hr_Ctrl_Rep3


title: Case 17: micronodular thymoma

description: micronodular thymoma


title: 2hr_N4shRNA_Rep3


title: 0hr_p400_480_Rep3


title: 0hr_Tip60_300_Rep3


title: 0hr_p400_315_Rep3


title: Case 16: micronodular thymoma

description: micronodular thymoma


title: Case 14: micronodular thymoma

description: micronodular thymoma


title: Case 11: micronodular thymoma

description: micronodular thymoma


title: Case 15: micronodular thymoma

description: micronodular thymoma


title: Case 10: micronodular thymoma

description: micronodular thymoma


title: 0hr_Ctrl_Rep3


title: 0hr_Tip60_299_Rep3


title: Case 9: micronodular thymoma

description: micronodular thymoma


title: Case 13: micronodular thymoma

description: micronodular thymoma


title: Case 12: micronodular thymoma

description: micronodular thymoma


title: Case 7: micronodular thymoma

description: micronodular thymoma


title: 6hr_p400_315_Rep2


title: 2hr_p400_315_Rep3


title: 6hr_Tip60_300_Rep2


title: 6hr_Tip60_299_Rep2


title: Case 6: micronodular thymoma

description: micronodular thymoma


title: Case 5: atypical type A thymoma

description: atypical type A thymoma

source_name: atypical type A thymoma


title: Case 4: atypical type A thymoma

description: atypical type A thymoma


title: 6hr_Ctrl_Rep2


title: 6hr_N4shRNA_Rep2


title: Case 3: atypical type A thymoma

description: atypical type A thymoma


title: Case 2: atypical type A thymoma

description: atypical type A thymoma


title: 2hr_p400_480_Rep2


title: 2hr_p400_315_Rep2


title: 2hr_Tip60_300_Rep2


title: 0hr_N4shRNA_Rep3


title: 6hr_p400_380_Rep2


title: 2hr_N4shRNA_Rep2


title: 2hr_Tip60_299_Rep2


title: 2hr_Ctrl_Rep2


title: Case 8: micronodular thymoma

description: micronodular thymoma


title: 0hr_p400_315_Rep2


title: 0hr_N4shRNA_Rep2


title: Case 1: atypical type A thymoma

description: atypical type A thymoma


title: 0hr_Ctrl_Rep2


title: 6hr_p400_480_Rep1


title: 2029_cpsf5cpsf6_rep2

city: Hanover

first: Christina

middle: M

last: Kelliher

state: NH

address: 7400 Hinman

department: Molecular & Systems Biology

institute: Geisel School of Medicine at Dartmouth

data_processing:
Confirm read quality: fastqc --format=fastq ./*.fastq.gz
Align reads: STAR --runThreadN 1 --runMode alignReads --genomeDir /N_crassa_NC12/STAR_genomebuild/ --sjdbGTFfile /N_crassa_NC12/FungiDB-45_NcrassaOR74A_3UTRs_7793_final.gff --sjdbGTFtagExonParentTranscript Parent --sjdbGTFfeatureExon three_prime_UTR --readFilesIn ./*.fastq --outFilterType BySJout --alignIntronMin 10 --alignIntronMax 3000 --outFileNamePrefix ./STAR_out/ --outSAMstrandField intronMotif --outFilterIntronMotifs RemoveNoncanonical
Count reads assigned to transcripts: htseq-count --format=bam --order=pos --stranded=reverse --mode=intersection-nonempty --type=three_prime_UTR --idattr=ID Aligned.out.sorted.bam /N_crassa_NC12/FungiDB-45_NcrassaOR74A_3UTRs_7793_final.gff > counts.txt
Normalize counts between 4 samples: custom R scripts available at https://github.com/cmk35
Output: Normalized gene counts used for analysis
Assembly: Neurospora crassa OR74A NC12 genome, FungiDB version 45, accessed 10/25/19: https://fungidb.org/fungidb/app/downloads/
Supplementary files format and content: tab-delimited text file includes Normalized Count values for each Sample

organism: Neurospora crassa

source_name: 2029: cpsf5/NCU09014::hphR cpsf6/NCU02152::hphR csr-1::cbox-luciferase::barR

growth_protocol: Neurospora conidia were inoculated onto cellophane-covered plates (1X Vogel’s Salts, 1.5% Noble agar w/v, 50 ng/ml biotin, 0.25% w/v glucose, 0.17% w/v arginine), and grown for 72 hours, 25C, constant light.

extract_protocol:
Neurospora tissue was isolated off the cellophane plate surface and frozen in liquid nitrogen. Frozen tissue was ground under liquid nitrogen using a mortar and pestle. Total RNA was extracted using Trizol (Invitrogen) and the Direct-zol RNA MicroPrep kit (Zymo).
Dartmouth Genomics procedures: total RNA QC pass, Lexogen QuantSeq 3’ REV library preparation kit, Illumina Mini-Seq sequencing, 75 bp stranded SE reads returned for each sample.


title: 6hr_p400_315_Rep1


title: 1785-1_control_rep2

source_name: 1785-1: csr-1::cbox-luciferase::barR MAT A


title: 6hr_Tip60_299_Rep1


title: 0hr_p400_480_Rep2


title: 0hr_Tip60_300_Rep2


title: 6hr_Ctrl_Rep1


title: 2029_cpsf5cpsf6_rep1


title: 6hr_N4shRNA_Rep1


title: 0hr_Tip60_299_Rep2


title: 1785-1_control_rep1


title: 2hr_p400_480_Rep1


title: 6hr_Tip60_300_Rep1


title: 2hr_Tip60_300_Rep1


title: 2hr_p400_315_Rep1


title: HiC_3D7_PfSETvs-KO_10-16h_rep2

city: Shanghai

first: Xuan

last: Cao

address: 320 Yueyang Road

institute: Shanghai Institute of Nutrition and Health,  Chinese Academy of Sciences

data_processing:
Raw sequencing reads were aligned to P. falciparum 3D7 reference genome (PlasmoDB release 43) , processed and ICE normalized using HiCPro package (version 2.11.4)
To eliminate the possible effects on data analyses of variable sequencing depths, we pooled valid read pairs of biological replicates and then normalized each sample to contain the same number of valid read pairs for downstream analyses.
For examination of global contact patterns, valid read pairs were then aggregated into contact matrix at 10kb resolution and normalized using ICE method to correct for experimental and technical biases.
We used GotHiC at 2kb resolution to identify long-range chromatin interactions. Interactions with pairs number >=5 and pvalue <= 0.1 were considered as high-confidence interactions. Additionally, intra-chromosomal interactions that linear distance <= 10kb were filtered out
Genome_build: P. falciparum 3D7 (PlasmoDB release 43)
Supplementary_files_format_and_content: ice matrix

organism: Plasmodium falciparum 3D7

source_name: PfSETvs-KO P. falciparum

growth_protocol: All parasites were cultured in an atmosphere consisting of 5% CO2, 5% O2 and 90% N2. Highly synchronous cultures of ring-stage parasites were used for the study.

extract_protocol: For Hi-C study, parasites were crosslinked by 1% formaldehyde for 10 min at room temperature followed by addition of 0.125 M glycine to stop crosslink. In situ Hi-C was performed as previously described with moderate modifications. Briefly, fixed parasites were lysed in 5 ml ice-cold Hi-C lysis buffer (10 mM Tris-HCl pH 7.5, 10 mM NaCl, 0.2% NP-40, 1X protease inhibitors) and rotated at 4 ℃ for 1 h. Pelleted nuclei were resuspended with 300 μl H2O, 44 μl of 10×NEBuffer 2 and 38 μl of 1% SDS and incubated at 65 °C for 10 minutes. 44 μL of 10% Triton X-100 were added and samples were rotated at 37 ℃ for 60 minutes to quench the SDS. To fragment DNA, 350 U endonucleases DpnII (NEB, R0543M) was added to the samples and samples were incubated at 37 ℃ overnight. Digested DNA was filled in the restriction fragment overhangs with Klenow (NEB, M0210L) and the DNA ends was marked with biotin. Chromatin was then ligated with T4 DNA ligase (thermo, EL0013) for 4 h at 16  ℃. DNAs were sonicated to reduce their size to 300-500 bp and immobilized on Dynabeads M-280 Streptavidin (Invitrogen,11205D).The process of ends repair, A-tailing and adaptor ligation referred to ChIP-seq method, the Hi-C library was amplified for 12-13 cycles and the PCR product was loaded on 1% agarose and size selected the 300-600bp band for sequencing. Then, Hi-C libraries were sequenced to generate at least 45 million paired-end reads (150 bp in length) per each donor/condition in a Novaseq 6000 platform (Illumina).

treatment_protocol: Parasites were synchronized to ring stage by 5% D-sorbitol treatment at 37℃ for 10 min at least one asexual cycles prior to samle collection.

tag: Stage

value: ring

tag: genotype/variation

value: PfSETvs-KO


title: HiC_3D7_PfSETvs-KO_10-16h_rep1


title: 2hr_Tip60_299_Rep1


title: 2hr_N4shRNA_Rep1


title: 2hr_Ctrl_Rep1


title: 0hr_p400_315_Rep1


title: HiC_3D7_wild-type_10-16h_rep2

source_name: wild-type P. falciparum

value: wild-type


title: 0hr_p400_480_Rep1


title: 0hr_Tip60_299_Rep1


title: RNAseq_3D7_PfSETvs-KO_10-16h_rep2

data_processing:
Raw read were aligned to P. falciparum 3D7 reference genome (PlasmoDB release 43) by using hisat2 (v.2.1.10) with parameters “--rna-strandness RF --dta”.
Mapped paired-end reads were firstly separated into forward and reversed strands according to the flag (forward FLAG83, FLAG163; reversed FLAG99, FLAG147) in sam files. FPKM (Fragments per kilobase per million mapped fragments) was calculated using stringtie (v2.1.4 ) with default parameters. Reads were counted for all genes by using HTseq-count.
Genome_build: P. falciparum 3D7 (PlasmoDB release 43)

extract_protocol: Cultured parasites were immediately pelleted and resuspended in TRIzol (ThermoFisher, Cat.N: 15596026). The samples were then processed using the RNeasy Mini Kit (QIAGEN, Cat.N: 74104) with DNase treatment, according to the manufacturer’s instructions. Purified RNA was then used for library generation using the NEBNext UltraTM Directional RNA Library Prep Kit following the manufacturer’s instructions.


title: EMRA-38

city: Lebanon

first: Lucas

middle: A.

last: Salas

address: 1 Medical Center Dr, DHMC

department: Epidemiology

description:
205624860032_R07C01
TEMRA

hyb_protocol: bisulphite converted DNA was amplified, fragmented and hybridised to Illumina Infinium Human MethylationEPIC Beadchip using standard Illumina protocol

scan_protocol: Arrays were imaged using BeadArray Reader using standard recommended Illumina scanner setting

data_processing:
BeadStudio software v3.2
matrix processed value definition: SeSAMe Processed Average Beta
matrix signal intensities value definition: Unmethylated and methylated signal intensities

label: Cy5 and Cy3

source_name: isolated CD8 terminally differentiated effector memory T;  leukoreduction filter 38

label_protocol: Standard Illumina Protocol

extract_protocol: genomic DNA was extracted and purified from LBC samples using Qiagen DNeasy Kit according to standard instructions

tag: age

tag: purity

tag: ethnicity

tag: arraytypes

value: IlluminaHumanMethylationEPIC


title: HiC_3D7_wild-type_10-16h_rep1

tag: strain


title: RNAseq_3D7_PfSETvs-KO_10-16h_rep1


title: EMRA-37

description:
205689130135_R08C01
TEMRA

source_name: isolated CD8 terminally differentiated effector memory T;  leukoreduction filter 37


title: EMRA-36

description:
205624860032_R05C01
TEMRA

source_name: isolated CD8 terminally differentiated effector memory T;  leukoreduction filter 36


title: 0hr_N4shRNA_Rep1


title: 0hr_Tip60_300_Rep1


title: EMRA-30

description:
205624860111_R07C01
TEMRA

source_name: isolated CD8 terminally differentiated effector memory T;  leukoreduction filter 30


title: 0hr_Ctrl_Rep1


title: Tip60_H_delcre

description: Single nucleus RNA-seq (10X) from deltacre infected hippocampal nuclei

data_processing:
FASTQ files were created using the standard bcl2fastq pipeline from Illumina. FASTQ files were concatenated across flowcell lanes and and across four sequencing runs.
Gene expression tables for each nuclear barcode were generated via the CellRanger 3.0.0 pipeline as designed by 10X Genomics. Samples were demultiplexed, and all Npas4fl/fl or Tip60fl/fl samples were merged using the CellRanger aggr function using default parameters.
The datasets were loaded into R and analyzed using the Seurat (v3) and Monocle3 packages. Nuclei were removed from the dataset if they contained fewer than 500 detected genes, displayed more than 5% of reads mapping to mitochondrial genes, or had RNA counts detected at a level greater than 2 standard deviations higher than the mean value in their assigned cell type.
Assembly: mm10
Supplementary files format and content: h5 filtered feature barcode matrices generated by Cellranger 3.0

source_name: Hippocampus

extract_protocol:
To isolate hippocampal nuclei, we placed flash-frozen hippocampal tissue in 0.5mL of buffer HB (0.25 M sucrose, 25 mM KCl, 5 mM MgCl2, 20 mM Tricine-KOH, pH 7.8, 1 mM DTT, 0.15 mM spermine, 0.5 mM spermidine) and dounced 5X with a loose pestle and 10X with a tight pestle. 5% IGEPAL CA-630 (32 µL) was added prior to douncing with a tight pestle 5-8 more times and filtering through a 40-µm strainer into a 15 mL conical collection tube. Buffer HB (3.5 mL) and 5 mL working solution (50% iodixanol, 25 mM KCl, 5 mM MgCl2, 20 mM Tricine-KOH, pH 7.8, supplemented with protease inhibitors, DTT, spermine and spermidine) were added. Homogenized tissue was gently layered on top of 1 mL of 30% iodixanol on top of a layer of 1 mL of 40% iodixanol (diluted from working solution). Samples were centrifuged at 10,000g for 18 min and 70 µL of nuclei was collected from the 30/40% iodixanol interface.
SnRNA-seq was performed using the 10X Genomics Chromium Single Cell Kit (v3). Each reaction lane was loaded with up to 10,000 nuclei from one hippocampal hemisphere (infected with either Cre-mCherry or ΔCre-GFP) from one mouse. Subsequent steps for cDNA amplification and library preparation were conducted according to the manufacturer’s protocol (10X Genomics). Samples were sequenced using Illumina NextSeq500 with 28 bp (R1), 56 bp (R2), and 8 bp(index).
Single nucleus RNA-seq (10X)

treatment_protocol: Postnatal day 21 day mice were injected with AAVs expressing either CRE virus (to remove the protein of interest) or DELCRE ('called DEL'), a nuclease deficient CRE. Mice were sacrificed 10 days later. To induce synchronous neuronal activity, mice were injected with 12 mg/kg kainic acid (KA).  Hippocampal tissue was harvested 6 hours later.

value: 6 hour post kainate stim

value: Hippocampal nuclei


title: Tip60_H_cre

description: Single nucleus RNA-seq (10X) from cre infected hippocampal nuclei


title: RNAseq_3D7_wild-type_10-16h_rep1


title: RNAseq_3D7_wild-type_10-16h_rep2


title: Tip60_G_delcre


title: Tip60_B_cre


title: EMRA-29

description:
205689130106_R07C01
TEMRA

source_name: isolated CD8 terminally differentiated effector memory T;  leukoreduction filter 29


title: Npas4_7_cre


title: round_spermatids2 (RS2), MeRIP-seq

city: Yangling

first: Xiaoxu

last: Chen

address: Litai street

institute: Northwest A&F University

description:
m6A modification
Methylated_RNA_sites.mRNA

data_processing:
RNA-seq: After raw data process, high quality clean reads were aligned to the reference genome (susScr3) with hisat2 software (v2.0.4). Then, cuffdiff software (part of cufflinks) was used to get the gene level FPKM based on the Ensembl gtf gene annotation file. The differentially expressed mRNA were identified and used for further analysis.
MeRIP-seq: After quality controlled by Q30, 3’ adaptor and low quality reads were removed by cutadapt software (v1.9.3). Clean reads of all libraries were aligned to the reference genome (SusScr3) by Hisat2 software (v2.0.4). Methylated sites were identified by MACS software and further used for differentially methylation analysis. Gene ontology analysis were performed by the differentially methylated protein coding genes.
Genome_build: SusScr3
Supplementary_files_format_and_content: abundance measurements - Methylated_RNA_sites.mRNA.xlsx, mRNA Expression Profiling.xlsx

organism: Sus scrofa

source_name: germ cells

extract_protocol:
Fractions with high purity of spermatogonia, pachytene spermatocytes and round spermatids were resuspended with Trizol (Invitrogen, USA) and total RNA was extracted for further analysis.
RNA seq: The rRNAs were removed from total RNA by using NEBNext rRNA Depletion Kit (New England Biolabs, Inc., Massachusetts, USA) following the manufacturer’s instruction. NEBNext® Ultra™ II Directional RNA Library Prep Kit (New England Biolabs, Inc., Massachusetts,USA) was used to construct RNA libraries. Library quality were confirmed by BioAnalyzer 2100 system (Agilent Technologies, Inc., USA). Library sequencing was performed on an Illumina Hiseq 4000 with 150 bp paired end reads.
MeRIP-seq: In brief, total RNA was extracted by using TRIzol Reagent (Invitrogen, USA). Denaturing agarose electrophoresis was used for confirming RNA integrity. Then, mRNA was isolated from total RNA by using Seq-Star T M poly(A) mRNA Isolation Kit (Arraystar, MD, USA). The m6A RNA immunoprecipitation was conducted by using GenSeq™ m6A RNA IP Kit (GenSeq Inc., China) according to the manufacturer’s instructions. Libraries were constructed from the samples with and without m6A immunoprecipitation by using NEBNext® Ultra II Directional RNA Library Prep Kit (New England Biolabs, Inc., MA, USA). Library was evaluated by the BioAnalyzer 2100 system (Agilent Technologies Inc., CA, USA) and sequenced on an Illumina Hiseq 4000 with 150 bp paired-end reads.  After quality controlled by Q30, 3’ adaptor and low quality reads were removed by cutadapt software (v1.9.3). Clean reads of all libraries were aligned to the reference genome (SusScr3) by Hisat2 software (v2.0.4). Methylated sites were identified by MACS software and further used for differentially methylation analysis. Gene ontology analysis were performed by the differentially methylated protein coding genes.

tag: cell type

value: round spermatids


title: CD8nv-37

description:
205689130106_R02C01
CD8nv

source_name: isolated CD8 naive; leukoreduction filter 37


title: Tip60_B_delcre


title: Tip60_G_cre


title: Npas4_6_cre


title: CD8nv-41

description:
205689130106_R08C01
CD8nv

source_name: isolated CD8 naive; leukoreduction filter 41