title: GIST-T1 AP H3K4me3

city: Sapporo

first: Hiromu

last: Suzuki

state: Hokkaido

address: S1, W17, Chuo-ku

department: Department of Molecular Biology

institute: Sapporo Medical University

description: GIST-T1 cells treated with 5-aza-dC plus 4-PBA

data_processing:
Basecalls performed using SETS version 4.
ChIP-seq reads were aligned to the hg38 genome assembly using Bowtie 2.
Peaks were called using MACS2.
Genome_build: hg38
Supplementary_files_format_and_content: BAM

source_name: GIST-T1 cells

growth_protocol: GIST-T1 cells were maintained in DMEM supplemented with 10% fatal bovine serum.

extract_protocol:
Lysates were clarified from sonicated nuclei and histone-DNA complexes were isolated with antibody.
Library construction was performed according to the manufacturer's instructions. Briefly, 100 ng of input DNA or the immunoprecipitate was ligated to adapters and PCR amplified using a SOLiD Fragment Library Construction kit (Applied Biosystems). Template bead preparation was carried out using a SOLiD ePCR kit V2 (Applied Biosystems) and a SOLiD Bead Enrichment kit (Applied Biosystems).

treatment_protocol: GIST-T1 cells were treated with 2 μM 5-aza-2’-deoxycytidine (5-aza-dC; Sigma-Aldrich) for 72 hours and then treated with or without 3 mM 4-phenylbutyric acid (4-PBA; Sigma-Aldrich) for an additional 48 hour, replacing drug and medium every 24 hours.

value: GIST

tag: chip-antibody

value: H3K4me3 (Upstate, clone MC315)

tag: treatment

value: 5-aza-dC + 4-PBA


title: GIST-T1 H3K4me3

status: Public on Jun 21 2021

type: SRA

country: Japan

description: GIST-T1 cells

library_source: genomic

molecule: genomic DNA

organism: Homo sapiens

tag: cell type

value: untreated


title: SHP77-YAP (wt) rep2

city: Toronto

first: Rod

last: Bremner

state: ON

address: 600 University Ave

department: Lunenfeld-Tanenbaum Research Institute

country: Canada

institute: Mount Sinai Hospital

description:
2x100 paired-end sequence reads
SHP77_YAP_4
processed data file: gene.counts_SHP77_Y79.matrix

library_source: transcriptomic

data_processing:
Illumina HiSeq Control Software (Y79 and SHP77 cells) or NextSeq Control Software (WERI, H209 and H2171 cells) was used for real-time base calling. The real-time base call (.bcl) files were converted to fastq files using CASAVA 1.8.2 (Y79 and SHP77) or Illumina bcl2fastq2 conversion software v2.17 (WERI, H209 and H2171).
Reads were adaptor trimmed. Read quality was assessed by FastQC v0.11.7 for each sample. Reads were processed for transcript quantification by using RSEM v1.3.0 with reference genome GRCh37.p13, and the resulting gene expression, as a matrix of raw counts.
Genome_build: GRCh37.p13
Supplementary_files_format_and_content: *.matrix: Tab-delimited text files with read raw counts for each gene and sample, with genes annotated by gene_id and gene_name.

molecule: total RNA

source_name: SCLC cell line

growth_protocol: Cell lines were cultured by standard protocols as outlined by the ATCC.

extract_protocol:
Total RNA was extracted using the Qiagen RNeasy mini kit.
RNA was accessed by Fragment Analyzer, and high quality total RNA was subjected to library preparation using the Illumina TruSeq Stranded mRNA Library Preparation Kit according to the manufacturer’s instructions.

treatment_protocol: Cells were harvested five days after transduction. Cell pellets were flash frozen and stored at -80°C until extraction.

tag: cell line

tag: genotype/variation

value: expressing wild-type human YAP


title: SHP77-YAP (S94A) rep1

description:
2x100 paired-end sequence reads
SHP77_S94A_3
processed data file: gene.counts_SHP77_Y79.matrix

value: expressing S94A mutant human YAP


title: H446 BAS_10uM

city: guangzhou

first: zhang

middle: zhen

last: hua

state: guangdong

address: xiaoguwei

department: college of pharmacy

country: China

institute: Sun Yat-Sen University

data_processing:
First, we used Trim Galore to automate quality and adapter trimming as well as quality control for FastQ files.
Then, the FASTQ-formatted sequence data were analyzed using a standard BWA-Bowtie2-Cufflinks workflow.
the Cufflinks package was applied for transcript assembly, quantification of normalized gene and isoform expression in RPKM (Reads per kilobase per million mapped reads) or FPKM (fragments per kilobase of exon per million fragments mapped),
and testing for differential expression (Cuffdiff).
Genome_build: hg19
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample

source_name: H446 cells cultured in the presence of BAS (10 uM)

growth_protocol: H446 cells were cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin . Cells were grown at 37℃ in 5% CO2 incubators.

extract_protocol:
RNA-seq libraries were from 1 μg total RNA which was extracted using the total RNA isolation kit (Tiangen, BJ, China) and reverse transcribed with reverse transcriptase M-MLV (Takara, DL, China), according to the manufacturer's instructions. Libraries were validated with an Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA). Sequencing was performed on an MGISEQ2000 SE50 sequencer at BGI Tech (Wu Han, China).
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: H446 cells were treated with vehicle or the small molecule BAS (5 μM) and BAS (10 μM), for 48 h before RNA extraction

value: H446 small cell lung cancer

value: 10 uM 3-Bromoascochlorin (BAS)


title: SHP77-YAP (S94A) rep2

description:
2x100 paired-end sequence reads
SHP77_S94A_4
processed data file: gene.counts_SHP77_Y79.matrix


title: Y79-YAP (S94A) rep2

description:
2x100 paired-end sequence reads
Y79_S94A_4
processed data file: gene.counts_SHP77_Y79.matrix

source_name: retinoblastoma cell line


title: H446 BAS_5uM

source_name: H446 cells cultured in the presence of BAS (5 uM)

value: 5 uM 3-Bromoascochlorin (BAS)


title: Y79-YAP (wt) rep2

description:
2x100 paired-end sequence reads
Y79_YAP_4
processed data file: gene.counts_SHP77_Y79.matrix


title: H446  Vehicle

source_name: H446 cells cultured in medium with vehicle control.

value: vehicle control


title: SHP77-Empty rep1

description:
2x100 paired-end sequence reads
SHP77_Empty_3
processed data file: gene.counts_SHP77_Y79.matrix

value: expressing Empty vector


title: Y79-YAP (S94A) rep1

description:
2x100 paired-end sequence reads
Y79_S94A_3
processed data file: gene.counts_SHP77_Y79.matrix


title: SHP77-Empty rep2

description:
2x100 paired-end sequence reads
SHP77_Empty_4
processed data file: gene.counts_SHP77_Y79.matrix


title: Y79-YAP (wt) rep1

description:
2x100 paired-end sequence reads
Y79_YAP_3
processed data file: gene.counts_SHP77_Y79.matrix


title: Line 9 rep.2 periderm

city: Girona

first: Olga

last: Serra

state: Girona

address: C/ Maria Aurèlia Capmany, 40

department: Biologia

country: Spain

institute: Universitat de Girona

description: Line 9 rep.2

data_processing:
RNA-seq data were first analysed for quality control with FastQC v0.11.2
The reads were aligned with Bowtie 2 (Langmead and Salzberg, 2012) against the S. tuberosum Group Phureja transcriptome generated from the genome assembly v4.03 (Potato Genome Sequencing Consortium et al., (2011) using the most recent version of the GFF3 genome annotation file (PGSC_DM_V403_genes.gff.zip) with the program gffread from the Cufflinks package
The quantification of transcript abundance was performed with eXpress 1.5.1
The eff_counts obtained from eXpress were passed as input to baySeq (Hardcastle et al., 2010) for the differential expression analysis. Transcripts with a likelihood ≥ 0.8 in the specified model and a False Discovery Rate (FDR) lower than 0.05 were considered as differentially expressed
Genome_build: PGSC v4.03 pseudomolecule
Supplementary_files_format_and_content: Matrix table with normalized gene counts for every gene and every sample

organism: Solanum tuberosum

source_name: potato tuber skin (periderm)

growth_protocol: in soil under long-day conditions (12 h light/12 h dark)

extract_protocol:
Total RNA was purified by means of the PureLink® Plant RNA Reagent (Ambion) using a modification of the standard protocol by repeating twice the step four and five of the protocol and adding a KOAc 2M (pH 5.5) precipitation step to remove polysaccharides. Final RNA precipitation was performed with the GlycoBlue™ Coprecipitant (Ambion). Genomic DNA was removed using TURBO DNA-free kit (Ambion) according to the manufacturer’s instructions.
The cDNA libraries were prepared using the TruSeq RNA Library Prep Kit (Illumina)

tag: tissue

value: tuber periderm

tag: genotype

value: StRIK-RNAi transgenic line 9

tag: growing conditions


title: Y79-Empty rep1

description:
2x100 paired-end sequence reads
Y79_Empty_3
processed data file: gene.counts_SHP77_Y79.matrix


title: Y79-Empty rep2

description:
2x100 paired-end sequence reads
Y79_Empty_4
processed data file: gene.counts_SHP77_Y79.matrix


title: case6_Substantia_Nigra_compacta_rep1

city: Stockholm

first: Eva

last: Hedlund

address: Svante Arrhenius v. 16C

department: Department of Biochemistry and Biophysics

country: Sweden

institute: Stockholm University

data_processing:
Base-calling using Illumina Cassava 1.7
Alignment to the human genome (hg38) performed using STAR with default parameters. RPKM were estimated using ¨rpkmforgenes¨ (Ramsköld et al., 2009).
For cases having more than 1 replicate per group, corresponding samples were averaged before the analysis so that each Case have only 1 SNc and 1 VTA.
Differential gene expression analysis was performed on the read counts using DESeq version 1.16.1. Only genes with reads in at least one third of the samples or more were considered in the analysis.
Genome_build: hg38
Supplementary_files_format_and_content: One processed data file is added providing abundance measurements (RPKM) organized as a matrix for all samples (tab-delimited txt).

molecule: polyA RNA

source_name: midbrain dopamine neurons

extract_protocol:
Lysis buffer is directly added to the cells after laser capture. No RNA extraction is conducted. The cell lysate is direcly used for library preparation.
Libraries for RNA-seq were prepared by using SMART-seq2 combined with Tn5 tagementation.

tag: case number (refer to table in paper)

tag: disease status

value: Control

tag: neurons

value: SNc


title: case4_Ventral_Tegmental_area

value: VTA


title: case5_Ventral_Tegmental_area


title: case18_Ventral_Tegmental_area


title: case4_Substantia_Nigra_compacta


title: case3_Substantia_Nigra_compacta_rep1


title: case18_Substantia_Nigra_compacta_rep2


title: SHP77-YAP (wt) rep1

description:
2x100 paired-end sequence reads
SHP77_YAP_3
processed data file: gene.counts_SHP77_Y79.matrix


title: case17_Ventral_Tegmental_area_rep2


title: case18_Substantia_Nigra_compacta_rep1


title: GIST-T1 input DNA

description: GIST-T1 cells


title: case17_Substantia_Nigra_compacta_rep3


title: case17_Substantia_Nigra_compacta_rep1


title: case15_Substantia_Nigra_compacta


title: case15_Ventral_Tegmental_area


title: case13_Substantia_Nigra_compacta


title: case17_Substantia_Nigra_compacta_rep5


title: case12_Substantia_Nigra_compacta_rep1


title: case17_Substantia_Nigra_compacta_rep2


title: case11_Substantia_Nigra_compacta


title: case12_Substantia_Nigra_compacta_rep2


title: case17_Substantia_Nigra_compacta_rep4


title: case3_Ventral_Tegmental_area_rep1


title: AF00115

city: Los Angeles

first: Miguel

last: Quintero

country: USA

institute: Cedars Sinai medical center

data_processing:
The circulating e-miRNA expression levels of the baseline blood samples of the  matched subjects were measured and processed through the workflow mentioned above
cut-off value of 500  was set as a minimal valid read counts per miRNA to eliminate the noise that had been produced in sequencing.  Expression data were further normalized using the DESeq method
Principal component analysis (PCA) was performed to reduce data dimensionality.
The principal components  (PCs) accounting for 95% of the total variance of the data were selected and calculated for each patient
The association between the PCs and recurrent ischemic events were evaluated via Mann-Whitney tests with  Bonferroni correction to determine the predictive PC for recurrent ischemic events and reduce the false discovery  rate; binary logistic model was then generated to calculate the sensitivity and specificity via a receiver operative  curve (ROC).
To further scale the contribution of each e-miRNA to the predictive PC, the absolute coefficients  of all the e-miRNAs in the PC formula were distributed in a quartile box plot, and the e-miRNAs with the coefficients  above 75% quartile were selected.
To interpret the regulatory effect of these selected e-miRNAs, functional  analysis and miRNA target analysis was carried out in Ingenuity Pathway Analysis (IPA) environment to mine  the significantly associated biological functions and downstream molecules
The up- and down-regulation of the  specific e-miRNA expression profile in the non-responders on the relevant biological functions and molecules  were predicted via a build-in Molecular Activity Predictor (MAP) tool
The networks, functional analyses, and  miRNA target analyses were generated through the use of IPA (QIAGEN Inc, https://www.qiagenbio-informatics.  com/products/ingenuity-pathway-analysis). A sensitivity analysis was conducted to address the potential impact  of including TIA in the primary endpoint
The expression levels of the relevant e-miRNAs, identified as described  above, were compared between the whole cohort and including exclusively subjects with stroke.
The statistical analyses including PCA and binary logistic regression analysis were performed in JMP (version  13). A two-tailed p value of <0.05 was considered statistically significant. The statistical significance of the  analysis in IPA environment was calculated by causal analytics algorithms which are based on a “master” network  derived from the Ingenuity Knowledge Base
Supplementary_files_format_and_content: Matrix table with raw gene counts for every miRNA and every sample for each group (responder to medical management and non responder)

source_name: Exosomal miRNA

extract_protocol:
Plasma sample with the volume of 500 μl was centrifuged at 1500 rpm for 5 min to remove residual cells and debris. The supernatant was transferred to a new 1.5 ml Eppendorf tube for exosome isolation. ExoQuick (Cat# EXOQ5A-1, System Biosciences, Palo Alto, CA) was added to the supernatant at 1:4 ratio (ExoQuick:Supernatant), mixed gently, and allowed to incubate for 30 min at 4 °C. After the incubation, the admixture was centrifuged at 1500 rpm for additional 30 min to recover the exosomes. Using ExoQuick-based precipitation techniques for exosome isolation and recovery is broadly recognized for better specific exosomal miRNAs recovery and lower intra-assay coefficients of variability (CVs) compared to other commercial exosome isolation kits, and is also backed in nearly 600 peer-review citations.
Exosomes extracted from the samples were processed for total RNA isola- tion using the SeraMir Exosome RNA Purification Column kit (Cat #RA808A-1, System Biosciences, Palo Alto, CA) according to the manufacturer’s instructions. For each sample, 1 μl of the final RNA eluate was used for meas- urement of small RNA concentration by Agilent Bioanalyzer Small RNA Assay using Bioanalyzer 2100 Expert instrument (Agilent Technologies, Santa Clara, CA). A small RNA concentration of 200 pg/ul was considers as a cutoff for subsequent library preparation.
Small RNA libraries were prepared for sequencing using standard Illumina protocols. Small Small RNA libraries were constructed with the CleanTag Small RNA Library Preparation Kit (Cat# L-3206, TriLink BioTechnologies, San Diego, CA) according to the manufacturer’s protocol
The final purified library was quanti- fied with High Sensitivity DNA Reagents (PO# G2933-85004, Agilent Technologies, Santa Clara, CA) and High Sensitivity DNA Chips (PO# 5067–4626, Agilent Technologies, Santa Clara, CA).
The libraries were pooled, and the 140 bp to 300 bp region was size selected on an 8% TBE gel (Ref# EC6215, Invitrogen by Life Technologies, Grand Island, NY). The size selected library was quantified with High Sensitivity DNA 1000 Screen Tape (PO #5067–5584, Agilent Technologies, Santa Clara, CA), High Sensitivity D1000 reagents (PO# 5067–5585, Agilent  Technologies, Santa Clara, CA), and the TailorMix HT1 qPCR assay (Cat# TM-505, SeqMatic, Union City, CA),  followed by a NextSeq High Output single-end sequencing run at SR75 using NextSeq 500/550 High Output v2  kit (Cat #FC-404-2005, Illumina, San Diego, CA) according to the manufacturer’s instructions.
Then, the open-source tools including FastqMcf, part  of the EA-utils package, and PRINSEQ were used to detect and remove N’s at the ends of reads, trim sequencing  adapters, and filter reads for quality and length. The improved set of sequence reads were mapped to the  reference genome using Bowtie and expression levels for exosomal miRNAs were calculated using R statistical  environment

tag: disease state

value: intracranial atherosclerosis

value: Serum

tag: phenotype

value: Non responder to medical management

tag: gender

value: Female

tag: age


title: case10_Substantia_Nigra_compacta_rep2


title: case10_Substantia_Nigra_compacta_rep1


title: case11_Ventral_Tegmental_area


title: case17_Ventral_Tegmental_area_rep3


title: 5capseq_SPT6_dTAG7_1h_1

city: Göttingen

first: Kristina

last: Žumer

address: Am Faßberg 11

country: Germany

institute: Max Planck Institute for Multidisciplinary Sciences

description: 5' capped RNA

data_processing:
Illumina software used for basecalling (Casava) and demultiplexing
MNase-seq: trimmed with Cutadapt (Martin, 2011) with options: -f fastq -q 30 -m 25 -O 13 -a AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC -A AGATCGGAAGAGCGTCGTGTAGGGAAAGAGTGTAGATCTCGGTGGTCGCCGTATCATT., followed by mapping to the human genome version GRCh38 merged with the D. melanogaster genome version Dm6 with the STAR mapper (Dobin et al., 2013) with options: --runThreadN 24 --readFilesCommand zcat --outFilterType Normal --alignEndsType Local --outFilterMatchNminOverLread 0.80 --outFilterMultimapNmax 1 --outFilterMismatchNmax 999 --outFilterMismatchNoverLmax 0.03 --alignMatesGapMax 716 --outSAMtype BAM SortedByCoordinate. Duplicates were marked with Picard tools (Broad-Institute). We then kept only uniquely mapped and properly paired reads for the downstream analyses. The counts from spike-ins were too low to be used for normalization, alternatively we used the standard normalization used for MNase-seq and generate normalization factors to correct for library sequencing depth from reads mapped to the human genome. These normalization factors were then used for the human gene counts and coverage profiles.
MNase-seq: mapped to the human genome version GRCh38 merged with the D. melanogaster genome version Dm6 with the STAR mapper (Dobin et al., 2013) with options: --runThreadN 24 --readFilesCommand zcat --outFilterType Normal --alignEndsType Local --outFilterMatchNminOverLread 0.80 --outFilterMultimapNmax 1 --outFilterMismatchNmax 999 --outFilterMismatchNoverLmax 0.03 --alignMatesGapMax 716 --outSAMtype BAM SortedByCoordinate.
MNase-seq: duplicates were marked with Picard tools (Broad-Institute). Uniquely mapped and properly paired reads were used for the downstream analyses. We obtained normalization factors to correct for library sequencing depth from reads mapped to the human genome.
MNase-seq: midpoints of the fragments were determined to obtain the positions of the putative nucleosome dyads. The dyad positions were then smoothed using normal kernel density smoothing with a bandwidth of 20 bp.
MNase-seq: the smoothed dyad coverage was then averaged per 5bp-bins to reduce the bigWig file size.
5cap-seq: first 8 nt of the reads containing the unique molecular identifier were first removed using UMI-tools and then trimmed with Cutadapt (Martin, 2011) with options: -f fastq -q 30 -m 25 -O 13 -a AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC
5cap-seq:  mapping to the human genome version GRCh38 merged with the D. melanogaster genome version Dm6 with the STAR mapper (Dobin et al., 2013) with options: --runThreadN 24 --readFilesCommand zcat --outFilterType BySJout --outFilterMultimapNmax 1 --alignEndsType Extend5pOfRead1 --alignSJoverhangMin 8 --alignSJDBoverhangMin 1 --outFilterMismatchNmax 999 --outFilterMismatchNoverLmax 0.05 --alignIntronMin 20 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --outSAMtype BAM SortedByCoordinate.
5cap-seq: dediplication of the mapped data with UMI-tools to remove any PCR duplicates.
5cap-seq: identify the first transcribed base (5’ end of the RNA) and used only this position in downstream analyses.
5cap-seq: the spike-in reads (processed as above) that mapped to the D. melanogaster genome were used to obtain normalization factors to normalize the human gene counts and coverage profiles.
Genome_build: GRCh38
Supplementary_files_format_and_content: bigwig files; normalized coverage generated in R with coverage function from the GenomicRanges R package (Bioconducor)

source_name: cell culture cells

growth_protocol: Cells were subcultured in RPMI medium (Thermo Fisher Scientific) supplemented with 10% fetal bovine serum (Thermo Fisher Scientific) and 1x GlutaMax (Thermo Fisher Scientific) and maintained at densities between 1x105 and 7x105 cells/ml

extract_protocol:
MNase-seq: the cells were washed with cold DPBS, resuspended in hypotonic buffer (10 mM HEPES pH 7.4, 2 mM MgCl2 and 0.5% NP40; supplemented with protease inhibitor cocktail (Sigma)) on ice and incubated 5 minutes. We added 200 gel units MNAse (NEB), heated up to 37°C, added CaCl2 to a final concentration of 2 mM and incubated at 37°C for 4 minutes before adding two volumes of 2xStop solution containing 20 mM Tris–HCl, pH 8, 4 mM MgCl2, 300 mM NaCl, 20 mM EDTA, 3 mM EGTA, 2% SDS, 0.2 μg/μL RNase A, and 2 pg/mL D.melanogaster mononucleosome spike-in DNA. The samples were then incubated at 37°C for 30 minutes, followed by addition of 200 µg proteinase K (Thermo Scientific) and 15 minutes of incubation at 55°C. The DNA fragments were then purified by phenol:chloroform:IAA extraction and precipitation with isopropanol.
5cap-seq: We spiked the samples with lysates containing 10 million D.melanogaster S2 cells and isolated RNA according to the manufacturers protocol (Qiazol).
MNase-seq: RNA libraries were prepared for sequencing using standard NEB protocols (NEBNext Ultra II DNA Library Prep Kit for Illumina).
5cap-seq: ligation of 5' DNA/RNA adaptor with an 8 nt unique molecular identifier, framgentation and random priming to generate the first strand, generation of second strand with biotinylated primer, caputure on strapavidin beads, ends-repair and A-tailing followed by adaptor ligation and amplification with primers with multiplexing barcodes.

treatment_protocol:
MNase-seq: 5 million cells were treated per biological replicate sample. We trated the cells with dTAG7 or control: DMSO for 1 or 4 hours.
5cap-seq: 5 million cells were treated per biological replicate sample. We trated the cells with dTAG7 or control: DMSO for 1 or 4 hours and then lysed the cell pellets in Qiazol (Qiagen)

value: chronic myelogenous leukemia

value: genome edited K562

tag: genome modification

value: SPT6 N-terminal dTAG

value: dTAG7 1h


title: 5capseq_SPT6_DMSO_4h_1

description: 5' capped RNA

value: DMSO 4h


title: 5capseq_SPT6_DMSO_4h_2

description: 5' capped RNA


title: 5capseq_SPT6_dTAG7_4h_1

description: 5' capped RNA

value: dTAG7 4h


title: 5capseq_SPT6_dTAG7_1h_2

description: 5' capped RNA


title: case17_Ventral_Tegmental_area_rep1


title: 5capseq_SPT6_DMSO_1h_1

description: 5' capped RNA

value: DMSO 1h


title: MNaseseq_SPT6_dTAG7_4h_2

description: MNase digested chromatin DNA


title: MNaseseq_SPT6_dTAG7_1h_1

description: MNase digested chromatin DNA


title: MNaseseq_SPT6_dTAG7_4h_1

description: MNase digested chromatin DNA


title: MNaseseq_SPT6_DMSO_4h_2

description: MNase digested chromatin DNA


title: MNaseseq_SPT6_DMSO_1h_1

description: MNase digested chromatin DNA


title: MNaseseq_SPT6_DMSO_4h_1

description: MNase digested chromatin DNA


title: MNaseseq_SPT6_DMSO_1h_2

description: MNase digested chromatin DNA


title: 5capseq_SPT6_DMSO_1h_2

description: 5' capped RNA


title: TTseq_SPT6_DMSO_4h_2

description: fragmented 4sU-labeled RNA

data_processing:
Library strategy: TT-seq
Illumina software used for basecalling (Casava) and demultiplexing
TT-seq: reads were trimmed with Cutadapt with options: -f fastq -q 30 -m 25 -O 13 -a AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC -A AGATCGGAAGAGCGTCGTGTAGGGAAAGAGTGTAGATCTCGGTGGTCGCCGTATCATT
TT-seq: the fastq files were mapped to the reference genome (GRCh38 + RNA spike-ins) using STAR aligner to generate the BAM alignment files with options: --runThreadN 24 --readFilesCommand zcat --outFilterType BySJout --outFilterMultimapNmax 1 --alignSJoverhangMin 8 --alignSJDBoverhangMin 1 --outFilterMismatchNmax 999 --outFilterMismatchNoverLmax 0.02 --alignIntronMin 20 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --outSAMtype BAM SortedByCoordinate
TT-seq: aligned duplicate reads were idnetified and marked with Picard tools (Broad Institute) and we then kept only uniquely mapped and properly paired reads for the downstream analyses.
TT-seq: normalization factors were generated counts from spike-ins counts with the rCube package (Bioconductor).
mNET-seq: reads were trimmed with Cutadapt with options: -f fastq -q 30 -m 20 -O 13 -a TGGAATTCTCGGGTGCCAAGGAACTCCAGTCAC -A GATCGTCGGACTGTAGAACTCTGAAC
mNET-seq: the fastq files were mapped to the reference genome (GRCh38 + sacCer3) using STAR aligner to generate the BAM alignment files with options: --runThreadN 24 --readFilesCommand zcat --outFilterType BySJout --outFilterMultimapNmax 1 --alignSJoverhangMin 8 --alignSJDBoverhangMin 1 --outFilterMismatchNmax 999 --outFilterMismatchNoverLmax 0.02 --alignIntronMin 20 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --outSAMtype BAM SortedByCoordinate
mNET-seq: aligned duplicate reads were idnetified and marked with Picard tools (Broad Institute) and we then kept only uniquely mapped and properly paired reads for the downstream analyses.
mNET-seq: normalization factors were generated counts from yeast gene counts or stably expressed genes with the DESeq2 package (Bioconductor).
Genome_build: GRCh38
Supplementary_files_format_and_content: bigwig files; normalized coverage generated in R with coverage function from the GenomicRanges R package (Bioconducor)

extract_protocol:
TT-seq: Six in vitro transcribed spike-in RNAs were added to the lysates (Qiazol, Qiagen) from 30 million cells, followed by extraction of RNA according to the manufacturer’s protocol.  This was followed by fragmentation by sonication and biotinylation of the 4sU-containing RNA. Two biotinylation reactions were set up per sample (150 µg fragmented total RNA, 10 mM Tris pH 7.5, 1 mM EDTA pH 8.0, 200 µg/ml HPDP-biotin, 40% DMSO) and then pooled after purification with the µMACS Streptavidin Kit (Miltenyi). The 4sU-labeled RNA was eluted by cleaving the HDPD-biotin with 100 mM DTT and then purified and concentrated with the miRNeasy Micro kit (Qiagen). 100 µg of the purified 4sU-labeled RNA was then used as input for the Ovation library prep kit (Nugen).
mNET-seq:  Cells were seeded the day before the experiment to get 100 million cells per sample the following day. We randomly assigned flasks for each treatment and treated cells with DMSO or dTAG7 ligand (final dTAG7 conc. 500 nM) for 1 or 4h, before extracting the chromatin-bound Pol II. The fractionation was done with 25 million cells per chromatin extraction. The cells were first washed with ice-cold DPBS, resuspended in 4 ml ice-cold HLB+N buffer (10 mM Tris-HCl (pH 7.5), 10 mM NaCl, 2.5 mM MgCl2 and 0.5% (vol/vol) NP-40) and incubated on ice for 5 minutes. The cell suspension was then underlaid with 1 ml HLB+NS buffer (10 mM Tris-HCl (pH 7.5), 10 mM NaCl, 2.5 mM MgCl2, 0.5% (vol/vol) NP-40 and 10% (wt/vol) sucrose) and centrifuged to pellet the nuclei at 0.44 x g. The supernatant and membrane debris was then removed and the nuclei were resuspended in 125 µl NUN1 lysis buffer (20 mM Tris-HCl (pH 7.9), 75 mM NaCl, 0.5 mM EDTA and 50% (vol/vol) glycerol) to which we added 1.2 ml NUN2 buffer to precipitate the chromatin (20 mM HEPES-KOH (pH 7.6), 300 mM NaCl, 0.2 mM EDTA, 7.5 mM MgCl2, 1% (vol/vol) NP-40 and 1 M urea) and incubated on ice for 15 minutes with occasional vortexing. The lysates are then centrifuged at ≥16,000g for 10 minutes at 4 °C to pellet the chromatin. These were then washed with MNase buffer and then digested with MNase (NEB) for 2 minutes at 37°C. The digestion was stopped by adding 5 µl of 500 mM EGTA and transferring onto ice. The reactions were then centrifuged at ≥16,000g for 5 minutes at 4 °C and then supernatant was diluted with 1 ml NET-2E buffer (50 mM Tris-HCl (pH 7.4, 150 mM NaCl,0.05% (vol/vol) NP-40 and 1 % Empigen BB) per fractionation and pooled per sample for the IP. We added 50 µl of antibody-conjugated beads (Dynabeads M-280 Sheep Anti-Mouse IgG rebound with 30 µg anti-CTD antibody) and incubated in the cold room for 1.5 h. This was followed by 6 washes with NET-2E buffer and phosphorylation of 5' RNA ends with T4 PNK 3’ phosphatase minus (NEB) by incubating the beads with phosphorylation buffer (1 x PNK Buffer, 0.1% Tween-20, 1.5 mM ATP, PNK) at 37°C for 6 min. We eluted the RNA with RNA extraction buffer (ZYMO Quick-RNA microPrep kit). We added 1 ng of fragmented S. cerevisae mRNA spike-in to the extraction buffer (1h dTAG7-treatment samples). In brief, these spike-ins were prepared from S. c. total RNA. Yeast cells were lysed in TSNTE buffer (2% triton x-100, 1% SDS, 100 mM NaCl, 10 mM Tris-HCl pH 8.0, 1 mM EDTA pH 8.0) and acidic phenol by several rounds of shaking with zirconia beads in the FastPrep machine, followed by RNA extraction and DNAse treatment (TURBO DNA-free DNAse, Thermo Fisher Scientific). Yeast mRNA was isolated with two rounds of purification with the Dynabeads mRNA Purification kit (Thermo Fisher Scientific). Yeast mRNA was then chemically fragmented with NEBNext Magnesium RNA Fragmentation Module (NEB) for 5 min at 94°C and the ends were repaired with T4 PNK (NEB) to be compatible for library input. Next, short immunoprecipitated RNA fragments were size selected (under 200 nt) and purified from the eluates (with or without spiked-in fragmented yeast mRNA) according to the manufacturers protocol (ZYMO Quick-RNA microPrep kit) and eluted in 7.5 µl nuclease-free water.
TT-seq RNA libraries were prepared for sequencing using standard Ovation library prep kit protocol (Nugen).
mNET-seq RNA libraries were prepared for sequencing using standard Illumina/NEB protocols (TruSeq Small RNA Library Prep Kit/NEBNext Small RNA Library Prep).

treatment_protocol:
TT-seq: 30 million cells were treated per biological replicate sample. Following the specified treatment (dTAG7 or control: DMSO for 1 or 4 hours), we supplemented the treatment medium with 500 µM 4-thiouridine (4sU) and metabolically labeled the cells for 5 min. The cells were pelleted by 2 min 2000 rpm centrifugation and lysed in Qiazol (Qiagen).
mNET-seq:  100 million cells were treated per biological replicate sample.We trated the cells with dTAG7 or control: DMSO for 1 or 4 hours.

tag: agent

value: DMSO

tag: time point


title: TTseq_SPT6_dTAG7_1h_2

description: fragmented 4sU-labeled RNA


title: TTseq_SPT6_dTAG7_1h_1

description: fragmented 4sU-labeled RNA


title: TTseq_SPT6_DMSO_1h_2

description: fragmented 4sU-labeled RNA


title: TTseq_SPT6_DMSO_4h_1

description: fragmented 4sU-labeled RNA


title: TTseq_RTF1_dTAG7_1h_B6

description: fragmented 4sU-labeled RNA

value: RTF1 N-terminal dTAG


title: TTseq_SPT6_dTAG7_4h_2

description: fragmented 4sU-labeled RNA


title: TTseq_SPT6_DMSO_1h_1

description: fragmented 4sU-labeled RNA


title: TTseq_RTF1_dTAG7_1h_H6

description: fragmented 4sU-labeled RNA


title: TTseq_SPT6_dTAG7_4h_1

description: fragmented 4sU-labeled RNA


title: TTseq_RTF1_DMSO_1h_H6

description: fragmented 4sU-labeled RNA


title: TTseq_PAF1_dTAG7_4h_2

description: fragmented 4sU-labeled RNA

value: PAF1 N-terminal dTAG


title: TTseq_RTF1_DMSO_1h_B6

description: fragmented 4sU-labeled RNA


title: MNaseseq_SPT6_dTAG7_1h_2

description: MNase digested chromatin DNA

tag: treatment


title: TTseq_CTR9_DMSO_4h_2

description: fragmented 4sU-labeled RNA

tag: cell type

value: CTR9 N-terminal dTAG


title: TTseq_PAF1_DMSO_4h_2

description: fragmented 4sU-labeled RNA

tag: cell line


title: TTseq_PAF1_dTAG7_4h_1

description: fragmented 4sU-labeled RNA


title: TTseq_CTR9_dTAG7_4h_2

description: fragmented 4sU-labeled RNA


title: TTseq_CTR9_DMSO_4h_1

description: fragmented 4sU-labeled RNA


title: TTseq_CTR9_dTAG7_4h_1

description: fragmented 4sU-labeled RNA


title: mNETseq_SPT6_dTAG7_1h_2

description: Pol II-associated RNA

data_processing:
Library strategy: mNET-seq
Illumina software used for basecalling (Casava) and demultiplexing
TT-seq: reads were trimmed with Cutadapt with options: -f fastq -q 30 -m 25 -O 13 -a AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC -A AGATCGGAAGAGCGTCGTGTAGGGAAAGAGTGTAGATCTCGGTGGTCGCCGTATCATT
TT-seq: the fastq files were mapped to the reference genome (GRCh38 + RNA spike-ins) using STAR aligner to generate the BAM alignment files with options: --runThreadN 24 --readFilesCommand zcat --outFilterType BySJout --outFilterMultimapNmax 1 --alignSJoverhangMin 8 --alignSJDBoverhangMin 1 --outFilterMismatchNmax 999 --outFilterMismatchNoverLmax 0.02 --alignIntronMin 20 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --outSAMtype BAM SortedByCoordinate
TT-seq: aligned duplicate reads were idnetified and marked with Picard tools (Broad Institute) and we then kept only uniquely mapped and properly paired reads for the downstream analyses.
TT-seq: normalization factors were generated counts from spike-ins counts with the rCube package (Bioconductor).
mNET-seq: reads were trimmed with Cutadapt with options: -f fastq -q 30 -m 20 -O 13 -a TGGAATTCTCGGGTGCCAAGGAACTCCAGTCAC -A GATCGTCGGACTGTAGAACTCTGAAC
mNET-seq: the fastq files were mapped to the reference genome (GRCh38 + sacCer3) using STAR aligner to generate the BAM alignment files with options: --runThreadN 24 --readFilesCommand zcat --outFilterType BySJout --outFilterMultimapNmax 1 --alignSJoverhangMin 8 --alignSJDBoverhangMin 1 --outFilterMismatchNmax 999 --outFilterMismatchNoverLmax 0.02 --alignIntronMin 20 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --outSAMtype BAM SortedByCoordinate
mNET-seq: aligned duplicate reads were idnetified and marked with Picard tools (Broad Institute) and we then kept only uniquely mapped and properly paired reads for the downstream analyses.
mNET-seq: normalization factors were generated counts from yeast gene counts or stably expressed genes with the DESeq2 package (Bioconductor).
Genome_build: GRCh38
Supplementary_files_format_and_content: bigwig files; normalized coverage generated in R with coverage function from the GenomicRanges R package (Bioconducor)


title: mNETseq_SPT6_dTAG7_1h_1

description: Pol II-associated RNA


title: BATF3-knockout clone #1

city: Vienna

first: Olaf

last: Merkel

address: Spitalgasse 23

department: Department of Pathology

country: Austria

institute: Medical University of Vienna

data_processing:
The raw fastq files were quality-checked using FastQC (v0.11.5) and low-quality ends (Phred score average of 4 consecutive bases <5) and adapters were removed using Trimmomatic (v0.36).
Alignment to the reference genome (GRCh38) with gene annotation (Ensembl v91) was performed using STAR (v2.5.3a). Post-processing was performed with Samtools (v1.4.1).
Gene counts were assessed by featureCounts (v1.5.2) and strand specificity of the sequencing was taken into account. Only uniquely aligned reads were considered. Overall statistics were continuously summarized by MultiQC (v1.4).
Differential expression calculation was performed in R (v3.4.4, http://www.R-project.org/) using DESeq2 (v1.16.1) / edgeR (v3.18.1).
Benjamini–Hochberg procedure was used to correct the P-values for multiple testing errors.
Genome_build: GRCh38, hg38
Supplementary_files_format_and_content: Matrix table with raw gene counts and normalized abundance measurements for all samples (txt files)

source_name: peripheral blood

growth_protocol: Cells were cultured in RPMI 1640 supplemented with FBS (10%) , penicillin (100 IU/ml) and streptomycin sulfate (50 mg/ml).

extract_protocol:
RNA isolation was performed using the RNeasy Mini Kit (Qiagen) according to the manufactuer's instructions with 1 μg total RNA used for the construction of sequencing library.
RNA-seq libraries were prepared using the NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (New England Biolabs) with Poly(A) selection as per the manufacturer’s instructions.

treatment_protocol: Cells were lentivirally transduced with either non-targeting (control) or BATF3 (knockout) sgRNA, followed by puromycin selection (1 μg/ml). Single clones were selected with mutations verified by Sanger sequencing and loss of protein confirmed by immunoblotting.

value: anaplastic large cell lymphoma cells

tag: alk status

value: ALK+

value: BATF3 knockout


title: mNETseq_SPT6_dTAG7_4h_2

description: Pol II-associated RNA


title: mNETseq_SPT6_DMSO_4h_2

description: Pol II-associated RNA


title: mNETseq_SPT6_DMSO_1h_2

description: Pol II-associated RNA


title: mNETseq_SPT6_dTAG7_4h_1

description: Pol II-associated RNA


title: mNETseq_SPT6_DMSO_4h_1

description: Pol II-associated RNA


title: mNETseq_RTF1_dTAG7_1h_H6

description: Pol II-associated RNA


title: mNETseq_SPT6_DMSO_1h_1

description: Pol II-associated RNA


title: mNETseq_RTF1_dTAG7_1h_B6

description: Pol II-associated RNA


title: mNETseq_RTF1_DMSO_1h_H6

description: Pol II-associated RNA


title: TTseq_PAF1_DMSO_4h_1

description: fragmented 4sU-labeled RNA


title: 3D culture 4T1 WT2

city: Bethesda

first: Li

last: Yang

state: Maryland

address: 37 Convent Dr.

department: NCI

institute: NIH

data_processing:
All analysis was performed using the CCBR pipeliner (https://github.com/CCBR/Pipeliner)
Genome_build: mm9
Supplementary_files_format_and_content: tab delimited files of DEGs from Limma analysis

organism: Mus musculus

growth_protocol:
Spheroids were made by growing cells in ultralow attachment plates for 3 days, then adding 75% matrigel and growing for 7 more days.
All cells were incubated in 37°C and 5% CO2. For 2D culture, cells were grown in 2% FBS.

extract_protocol: The Zymo Quick-DNA/RNA miniprep kit was used to extract RNA.

treatment_protocol: 4T1 iCasp9 Padi4+/+ and Padi4 -/- cells were induced with AP1903 to produce exsporosi and apoptotic debris respectively and then were added into cell cultures at a ratio of 5,000 cells to 500 recipient cells for the appropriate conditions.

tag: genotype/variation

value: WT


title: 2D culture 4T1 with vehicle2

value: vehicle


title: 2D culture 4T1 with vehicle3


title: 3D culture 4T1 KO3

value: KO


title: 3D culture 4T1 KO1


title: 2D culture 4T1 with exsporosi2

value: exsporosi


title: 2D culture 4T1 with exsporosi1


title: 3D culture 4T1 WT3


title: 2D culture 4T1 with exsporosi3


title: 2D culture 4T1 with apoptotic debris2

value: apoptotic debris


title: 2D culture 4T1 with apoptotic debris3


title: 2D culture 4T1 with apoptotic debris1


title: 3D culture 4T1 KO2


title: mNETseq_RTF1_DMSO_1h_B6

description: Pol II-associated RNA

value: chronic myelogenous leukemia

value: genome edited K562


title: 3D culture 4T1 WT1


title: 2D culture 4T1 with vehicle1


title: L8-P7_S187

city: Leuven

first: Bart

last: de Strooper

address: Campus Gasthuisberg, Herestraat 49, bus 602

department: VIB-KU Leuven Center for Brain & Disease Research

country: Belgium

institute: KULeuven

description:
L8-P13_S373
L8-P14_S374
L8-P15_S375
L8-P16_S376

data_processing:
To confirm quality of the raw reads, a FastQC analysis was performed. Reads were aligned to the mouse mm10 genome using STAR with default options. A count table was generated from the alignments using Subread/FeatureCounts 1.5.1.
The count matrix was analyzed with Seurat 2.3.1, using the following workflow: Cells with less than 200 genes or more than 6,000 genes (likely doublets) were removed, as well as cells whose mitochondrial gene expression accounted for more than 20% of the total gene expression of that cell. Genes found in less than 0.5% of cells were also excluded. Following this filtering, 709 of the original 768 cells remained (with a median read depth of 630,131 reads per cell and median of 2,646 genes per cell).
Data was log-normalized and scaled by a factor of 10,000.
The normalized count data was subsequently regressed on the number of reads.
The first 30 PCAs were used to identify clusters (using default parameters).
Cluster markers were determined by a differential expression analysis of each cluster versus all other clusters combined. Seurat’s “AddModuleScore” function, with default parameters, was used to calculate gene set scores of cell type-specific markers extracted from previous studies. Boxplots of these scores were used to assign each cluster to a cell type. We confirmed these assignments by comparing our cluster markers to unique markers of cell type defined by the aforementioned studies to ensure that there was no ambiguity in the assignment of cell types.
Gene differential expression between experimental (miR-132 overexpression) and control conditions (within each cell population) was performed. Differential expression (for analysis of cluster markers and for analysis between controls and experimental conditions) was performed as follows: p-values were calculated using a Wilcoxon rank-sum test (using Seurat’s “FindMarkers” function), and corrected for multiple testing using a Bonferroni correction. Log fold changes were calculated using the natural logarithm. Genes with adjusted p-values of < 0.05 were considered significantly differentially expressed.
Functional enrichment analysis was performed by using Ingenuity Pathway Analysis. By convention, the 150 transcripts with the most negative (‘Downregulated Transcripts’) or most positive (‘Upregulated Transcripts’) log fold changes after differential gene expression (comparing miR-132-overexpression to control samples) were selected per cluster for analysis.
Genome_build: mm10 genome (Ensembl Version 88)
Supplementary_files_format_and_content: Raw count matrix: neurogenesis_mirOE_raw_counts.tsv.gz

source_name: Dentate gyrus

extract_protocol:
For FACS isolation of single cells, four miR-132-injected and four control-injected Nestin:GFP male mice were used and 94 cells were initially sorted per mouse.
FACS-sorted cells were processed using the Smart-seq2 protocol. Briefly, RNA was reverse-transcribed using biotinylated oligo-dT primers and biotinylated template-switching (TSO) oligonucleotides and subsequently cDNA was preamplified using the KAPA HiFi Hot Start DNA Polymerase and biotinylated ISPCR primers. cDNA was purified using AMPURE XP Agencourt beads, concentration was calculated with the Quantifluor dsDNA kit and adjusted on an Echo 525 Liquid Handler. 100 pg cDNA per cell were used for library preparation with Nextera XT DNA Library Preparation Kit. Finally, following purification on magnetic beads the libraries were multiplexed and sequenced using the Illumina NextSeq500 platform.

treatment_protocol: For miR‐132 overexpression, 2 month-old Nestin:GFP mice received either a miR‐132 mimic or a negative control oligonucleotide in a 3 μl mix with lipofectamine 2000 (at a 1:1 volume ratio). Injections of 150 pmol oligo each were performed once a week for 5 weeks in total

tag: isolation

tag: mouse id

tag: date


title: L8-P9_S189

description:
L8-P21_S381
L8-P22_S382
L8-P23_S383
L8-P24_S384


title: L8-P2_S182

description:
L8-O17_S281
L8-O18_S282
L8-O19_S283
L8-O20_S284

value: Control


title: L8-P8_S188

description:
L8-P17_S377
L8-P18_S378
L8-P19_S379
L8-P20_S380


title: L8-P6_S186

description:
L8-P9_S189
L8-P10_S190
L8-P11_S191
L8-P12_S192


title: L8-P24_S384

description:
L8-O13_S277
L8-O14_S278
L8-O15_S279
L8-O16_S280


title: L8-P5_S185

description:
L8-P5_S185
L8-P6_S186
L8-P7_S187
L8-P8_S188


title: L8-P3_S183

description:
L8-O21_S285
L8-O22_S286
L8-O23_S287
L8-O24_S288


title: L8-P4_S184

description:
L8-P1_S181
L8-P2_S182
L8-P3_S183
L8-P4_S184


title: L8-P20_S380

description:
L8-N21_S369
L8-N22_S370
L8-N23_S371
L8-N24_S372


title: L8-P21_S381

description:
L8-O1_S85
L8-O2_S86
L8-O3_S87
L8-O4_S88


title: L8-P22_S382

description:
L8-O5_S89
L8-O6_S90
L8-O7_S91
L8-O8_S92


title: L8-P23_S383

description:
L8-O9_S93
L8-O10_S94
L8-O11_S95
L8-O12_S96


title: L8-P19_S379

description:
L8-N13_S361
L8-N14_S362
L8-N15_S363
L8-N16_S364


title: L8-P1_S181

description:
L8-N17_S365
L8-N18_S366
L8-N19_S367
L8-N20_S368


title: L8-P17_S377

description:
L8-N5_S173
L8-N6_S174
L8-N7_S175
L8-N8_S176


title: L8-P14_S374

description:
L8-M17_S269
L8-M18_S270
L8-M19_S271
L8-M20_S272


title: L8-P18_S378

description:
L8-N9_S177
L8-N10_S178
L8-N11_S179
L8-N12_S180


title: L8-P10_S190

description:
L8-M1_S73
L8-M2_S74
L8-M3_S75
L8-M4_S76


title: L8-P15_S375

description:
L8-M21_S273
L8-M22_S274
L8-M23_S275
L8-M24_S276


title: L8-P13_S373

description:
L8-M13_S265
L8-M14_S266
L8-M15_S267
L8-M16_S268


title: L8-P16_S376

description:
L8-N1_S169
L8-N2_S170
L8-N3_S171
L8-N4_S172


title: L8-P11_S191

description:
L8-M5_S77
L8-M6_S78
L8-M7_S79
L8-M8_S80


title: L8-O9_S93

description:
L8-L21_S357
L8-L22_S358
L8-L23_S359
L8-L24_S360


title: L8-P12_S192

description:
L8-M9_S81
L8-M10_S82
L8-M11_S83
L8-M12_S84


title: L8-O5_S89

description:
L8-L5_S161
L8-L6_S162
L8-L7_S163
L8-L8_S164


title: L8-O6_S90

description:
L8-L9_S165
L8-L10_S166
L8-L11_S167
L8-L12_S168


title: L8-O7_S91

description:
L8-L13_S349
L8-L14_S350
L8-L15_S351
L8-L16_S352


title: L8-O3_S87

description:
L8-K21_S261
L8-K22_S262
L8-K23_S263
L8-K24_S264


title: L8-O4_S88

description:
L8-L1_S157
L8-L2_S158
L8-L3_S159
L8-L4_S160


title: L8-O2_S86

description:
L8-K17_S257
L8-K18_S258
L8-K19_S259
L8-K20_S260


title: L8-O21_S285

description:
L8-K1_S61
L8-K2_S62
L8-K3_S63
L8-K4_S64


title: L8-O15_S279

description:
L8-I21_S249
L8-I22_S250
L8-I23_S251
L8-I24_S252


title: L8-O23_S287

description:
L8-K9_S69
L8-K10_S70
L8-K11_S71
L8-K12_S72


title: L8-O14_S278

description:
L8-I17_S245
L8-I18_S246
L8-I19_S247
L8-I20_S248


title: L8-O1_S85

description:
L8-J17_S341
L8-J18_S342
L8-J19_S343
L8-J20_S344


title: L8-O17_S281

description:
L8-J5_S149
L8-J6_S150
L8-J7_S151
L8-J8_S152


title: L8-O16_S280

description:
L8-J1_S145
L8-J2_S146
L8-J3_S147
L8-J4_S148


title: L8-O20_S284

description:
L8-J21_S345
L8-J22_S346
L8-J23_S347
L8-J24_S348


title: L8-O24_S288

description:
L8-K13_S253
L8-K14_S254
L8-K15_S255
L8-K16_S256


title: L8-O8_S92

description:
L8-L17_S353
L8-L18_S354
L8-L19_S355
L8-L20_S356


title: L8-O19_S283

description:
L8-J13_S337
L8-J14_S338
L8-J15_S339
L8-J16_S340


title: L8-O13_S277

description:
L8-I13_S241
L8-I14_S242
L8-I15_S243
L8-I16_S244


title: L8-O10_S94

description:
L8-I1_S49
L8-I2_S50
L8-I3_S51
L8-I4_S52


title: L8-O22_S286

description:
L8-K5_S65
L8-K6_S66
L8-K7_S67
L8-K8_S68


title: L8-O11_S95

description:
L8-I5_S53
L8-I6_S54
L8-I7_S55
L8-I8_S56


title: L8-N5_S173

description:
L8-H5_S137
L8-H6_S138
L8-H7_S139
L8-H8_S140


title: L8-N4_S172

description:
L8-H1_S133
L8-H2_S134
L8-H3_S135
L8-H4_S136


title: L8-O12_S96

description:
L8-I9_S57
L8-I10_S58
L8-I11_S59
L8-I12_S60


title: L8-N7_S175

description:
L8-H13_S325
L8-H14_S326
L8-H15_S327
L8-H16_S328


title: L8-N6_S174

description:
L8-H9_S141
L8-H10_S142
L8-H11_S143
L8-H12_S144


title: L8-N3_S171

description:
L8-G21_S237
L8-G22_S238
L8-G23_S239
L8-G24_S240


title: L8-N9_S177

description:
L8-H21_S333
L8-H22_S334
L8-H23_S335
L8-H24_S336


title: L8-N8_S176

description:
L8-H17_S329
L8-H18_S330
L8-H19_S331
L8-H20_S332


title: L8-N23_S371

description:
L8-G9_S45
L8-G10_S46
L8-G11_S47
L8-G12_S48


title: L8-N21_S369

description:
L8-G1_S37
L8-G2_S38
L8-G3_S39
L8-G4_S40


title: L8-N2_S170

description:
L8-G17_S233
L8-G18_S234
L8-G19_S235
L8-G20_S236


title: L8-N20_S368

description:
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description:
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title: miR_MSC_P12_REP3 [P12413]

city: Shenzhen

first: Ziming

last: Wang

state: Guangdong

address: 3025 Shennan Zhong Lu

department: Department of Orthopedics

institute: the eighth affiliated hospital of Sun Yat-sen University

data_processing:
After RNA integrity evaluation (Agilent Technologies, USA), rRNA remove, cDNA synthesis and PCR amplification, the sequencing database was built at BGI (Beijing Genomic Institute in Shenzhen) using the IlluminaHiSeq 3000 platform. Then the data was filtered with SOAPnuke (v1.5.2) to obtain clean reads.
The clean reads were obtained and stored in FASTQ format. HISAT2 (v2.0.4) and Bowtie2 (v2.2.5) were applied to align the clean reads to the reference gene set.
Expression level of gene was calculated by RSEM (v1.2.12).
Differential expression analysis was performed using DESeq2 (v1.4.5)15 with a Q value ≤ 0.05
Genome_build: Grch38
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

source_name: Bone marrow_miR_MSC_P12

growth_protocol: Cells were seeded in 12-well plates and cultured in Dulbecco’s modified Eagle’s medium (DMEM; Gibco) containing 10% fetal bovine serum (FBS; Gibco) at 37°C in a 5% CO2 atmosphere. The medium was replaced every 3 days.

extract_protocol:
MSCs total RNA samples of P4, P6 and P12 were separately treated with TRIzol (TAKARA).
The extracted total RNA was first removed using a biotin-labeled specific probe (Ribo-Zero™ rRNA Removal Kit) to remove the ribosomal rRNA.After purification, the RNA is fragmented at a certain temperature and ionic environment.A cDNA strand was then synthesized using a random primer and reverse transcriptase in the TruSeq® Stranded kit, and then a double-stranded cDNA was synthesized using DNA polymerase I and RNaseH.During cDNA two-strand synthesis, the RNA template is removed and dTTP is replaced by dUTP.The involvement of dUTP prevents the two strands of cDNA from being amplified in the subsequent process because the polymerase cannot cross the dUTP site on the template as it extends.The double-stranded cDNA product is then linked with an "A" base and connector.The ligation products will be amplified and purified to produce the final cDNA library.Finally, the constructed sequencing library was sequenced.
For miRNAs, Experiment Pipeline Steps for Small RNA Sequencing 1) Filter small RNA: separate 18-30nt RNA segment by PAGE gel. 2) 3' adaptor ligation: link 5-adenylated, 3-blocked single-stranded DNA adapter to 3'  end of selected small RNAs from step 1. 3) Reverse primer annealing: the RT primer is added to the solution from step 2, and is cross-linked to the 3' adapter of RNAs and excessive free 3' adapter. 4) 5' adaptor ligation: the 5' adaptor is linked to the 5' end of the product from step 3. The connection (5鈥 adaptor) will be attached to the 5鈥 end only, but it is not c onnected to the 3' adaptor or RT primer hybrid chain. This greatly reduces self-ligation.  5) Strand cDNA synthesis: reverse extend the RT primer in step 3 to synthetize strand cDNA. 6) PCR amplification: use high-ping polymerase to amplify cDNA, enrich cDNA with both 3' and 5' adaptor. 7) Library fragment selection: separate the PCR product of 100~120bp by PAGE gel to eliminate primer-dimer and other products.  8) Library quantitative and pooling cyclization.

treatment_protocol: MSCs were digested with 0.25% trypsin supplemented with 0.53mM EDTA (Gibco) to passaging cells to generation 4, 6, 12, respectively.

value: Bone marrow-derived mesenchymal stem cells

tag: generation

value: Generation 12

value: Expansion in vitro to passage 12


title: miR_MSC_P12_REP1 [P12393]

value: Generation 4


title: miR_MSC_P12_REP2 [P12411]

value: Generation 6


title: L7-E7_S223

description:
L8-D13_S301
L8-D14_S302
L8-D15_S303
L8-D16_S304


title: L7-E6_S222

description:
L8-D9_S117
L8-D10_S118
L8-D11_S119
L8-D12_S120


title: L7-F5_S125

description:
L8-H5_S137
L8-H6_S138
L8-H7_S139
L8-H8_S140


title: L7-F10_S130

description:
L8-E1_S25
L8-E2_S26
L8-E3_S27
L8-E4_S28


title: L7-E8_S224

description:
L8-D17_S305
L8-D18_S306
L8-D19_S307
L8-D20_S308


title: miR_MSC_P6_REP2 [P6411]

source_name: Bone marrow_miR_MSC_P6

value: Expansion in vitro to passage 6


title: miR_MSC_P6_REP3 [P6413]


title: L7-E4_S220

description:
L8-D1_S109
L8-D2_S110
L8-D3_S111
L8-D4_S112


title: L7-E5_S221

description:
L8-D5_S113
L8-D6_S114
L8-D7_S115
L8-D8_S116


title: miR_MSC_P6_REP1 [P6393]


title: L7-F20_S320

description:
L8-F21_S321
L8-F22_S322
L8-F23_S323
L8-F24_S324