title: ACS-1021 iPS-derived Cardiomyocytes 3

city: Jerusalem

first: Avner

last: Ehrlich

address: Silberman 3-512, Edmond J. Safra Campus (Givat Ram)

department: Bioengineering

country: Israel

institute: The Hebrew University of Jerusalem

data_processing:
RNA-seq reads for adult and fetal human cardiac tissues were downloaded from GSEA series GSE156707 along with accompanying metadata. AC16 data was downloaded from GSEA series GSE201076 along with accompanying metadata. RNA-seq reads for atrial muscle, ventricle muscle, SA node, and AV node were downloaded from GSEA series GSE203372 with accompanying metadata. Endocardium data was downloaded from GSE195911, epicardium was downloaded from GSE165450, and cardiac fibroblasts were downloaded from GSE167368. The reads were mapped to the UCSC human transcriptome (genome build hg19) using Bowtie2 . Expression levels of all genes were quantified using RSEM, yielding an expression matrix of inferred gene counts . Differential expression analysis was performed using R package DESeq2 using the default parameter . Reported are p-values from a negative binomial Wald-test.    In the endothelial marker analysis, The reads were mapped to the UCSC rat transcriptome (genome build rn6) using Bowtie2 . Expression levels of all genes were quantified using RSEM, yielding an expression matrix of inferred gene counts . Differential expression analysis was performed using R package DESeq2 using the default parameter . Isolated primary rat endothelial cells were downloaded from GSE133717.
Hierarchical clustering, heat maps, correlation plots, and similarity matrices were created in Morpheus. Gene ontology enrichment analyses and clustering were performed using DAVID Informatics Resources 6.7  and PANTHER Classification System . Metabolic network maps were created using McGill’s Network Analyst Tool using the KEGG database .
Assembly: genome build hg19; genome build rn6
Supplementary files format and content: tab-delimited text file includes raw counts for each sample

source_name: Cardiac

growth_protocol: Human induced pluripotent stem cell lines: UN-1 (healthy Middle Eastern male, 35 years old), ACS-1021 (healthy Caucasian male, 72 years old), and ACS-1028 (healthy African American female, 31 years old) were cultured on growth factor reduced Matrigel (BD Biosciences, San Jose, CA) in mTeSR-1 media (StemCell Technologies, Canada) with daily medium changes. Cells were obtained from the Technion (Prof. Lior Gepstein) and ATCC (Maryland, USA) respectively, authenticated at the source, and tested for mycoplasma contamination using PCR. Cells were serially passaged using 0.5X TrypLE Enzyme (Thermo Fisher Scientific, Waltham, MA, USA) supplemented with 0.5mM EDTA (Invitrogen, Thermo Fisher Scientific).  HEK 293T cells (ATCC, USA) were seeded in 10 cm cell culture plates at a density of 4x106 cells/plate. The cells were maintained in 293T medium composed of DMEM high glucose (4.5 g/l; Merck, USA) supplemented with 10% FBS (BI, Israel), 1x NEAA (BI, Israel), and 2 mM L-alanine-L-glutamine (BI, Israel).  Rat primary cardiac microvascular endothelial cells (CECs; Vec Technologies, USA) were cultured on gelatin-coated flasks using Endothelial Cell Growth Medium-2MV (Lonza, Switzerland) according to the manufacturer’s directions.  All cells were cultured under standard conditions in a humidified incubator at 37°C under 5% CO2. All cultured cells were tested routinely for mycoplasma contamination using PCR.

extract_protocol:
RNA was extracted in three separate experimental repeats from ACS-1021 hiPSC-derived cardiomyocyte (2D) and ACS-1021 cardiac organoid cultures using RNeasy Micro kit (Qiagen, USA) according to the manufacturer’s direction.
Library preparation and RNA sequencing were performed by the Hebrew University Center for Genomic Technologies. Briefly, library construction was conducted using Illumina TruSeq RNA Library Prep V2 Kit (Illumina, USA) and sequenced on Illumina NextSeq500 with single-end, 86 bp reads using the High Output V2 Kit.

treatment_protocol: Cardiac differentiation of human hiPSCs  The hiPSCs were seeded on Matrigel in mTeSR-1 medium and allowed to reach 80-90% confluence. Cardiac differentiation was conducted as previously described 90. Briefly, a basal medium named CDM3 was used (RPMI-1640, 500μg/mL recombinant human serum albumin, 213 μg/mL L-ascorbic acid 2-phosphate, and 1% penicillin/streptomycin). At day zero of differentiation, the medium was replaced with CDM3 supplemented with 6μmol/L CHIR99021 (Stemgent) for two days. On day two of the differentiation, the culture medium was switched to CDM3 medium supplemented with 2μM Wnt-C59 (Selleckchem), for an additional two days. From day four onward, cells were cultured with RPMI supplemented with B27 without Insulin; Gibco, USA). Generation of GFP-CECs Plasmid encoding GFP reporter upstream of a minimal CMV promoter was purchased from System Biosciences and validated in-house. Plasmids were acquired as bacterial LB-agar stabs and used per the manufacturer’s instructions. Briefly, each stab was first seeded into agar LB (Bacto Agar; BD, USA) in 10 cm plates. Then, single colonies were inoculated into flasks containing LB (BD Difco LB Broth, Lennox; BD, USA) and 100 µg/ml penicillin (BI, Israel). Transfection-grade plasmid DNA was isolated from each flask using the ZymoPURE II Plasmid Maxiprep Kit (Zymo Research, USA) according to the manufacturer’s instructions. HEK 293T cells were transfected with a GFP-expressing plasmid and the packaging plasmids using the TransIT-LT1 transfection reagent (Mirus Bio, USA) according to the provider’s instructions. Briefly, 6.65 µg GFP lentivector plasmid, 3.3µg pVSV-G, and 5µg psPAX2 were mixed in Opti-MEM reduced serum medium (Gibco, USA), with 45µl of TransIT-LT1, kept at room temperature to complex and then added to each plate. Following 18h of incubation, the transfection medium was replaced with 293T medium and virus-rich supernatant was harvested after 48h and 96h. The supernatant was clarified by centrifugation (500 x g, 5 min) and filtred (0.45 µm, Millex-HV, MerckMillipore). Packaging plasmids were a kind gift from the Nissim Benvenisti Lab, HUJI, Jerusalem, Israel.  Organoid Culture 4.5×106 hiPS-derived cardiomyocytes and 2.3×106 microvascular cardiac endothelial cells were counted and mixed in 100 µl of growth factor reduced Matrigel (BD Biosciences, San Jose, CA). A volume of 1.1 μl of the cell mixture was injected into each microwell and left to gel at 37C for 30 minutes. Cardiac organoid medium was composed of  RPMI-1640 supplemented with B27 minus insulin and 5 ng/ml Vascular Endothelial Growth Factor (VEGF-A, Peprotech). Cardiac organoid medium was added following gelation and changed every 48 hours. The cell mixture aggregates by day 4, start beating on day 10, and creates a smooth cardiac organoid with a synchronized beating by day 25 of seeding (Fig. 1A-B). The average organoid diameter was approximately 500 µm.

value: Cardiac

value: Cardiomyocytes


title: HEK293CDK7as_MNaseChIPseq_Input_MED26_TFIIE_TFIIB_NELF_1-NM-PP1_30min_Rep3

status: Public on Jun 01 2024

type: SRA

city: Goettingen

first: Michael

last: Lidschreiber

address: Am Fassberg 11

country: Germany

institute: Max-Planck-Institute for Multidisciplinary Scienes

library_source: genomic

data_processing:
All sequencing data were demultiplexed with Illumina bcl2fastq demultiplexing software, provided the barcodes for each of the samples, and subsequently quality checked with FastQC (Babraham-Institute) (Andrews, 2010).
TT-seq: For all samples, paired-end 75 base reads were mapped using STAR (v 2.6.1b) (Dobin et al., 2013) to the GRCh38.p13 genome assembly merged with synthetic RNA spike-in sequences (Schwalb et al., 2016; Żylicz et al., 2019) with a maximum 2% mismatches [mapping parameters: --outFilterMismatchNoverLmax 0.02 --outFilterMultimapScoreRange 0 --alignIntronMax 500000]. SAMtools (v 1.3.1) (Li et al., 2009) was used to quality filter SAM files, whereby alignments with MAPQ smaller than 7 (-q 7) were skipped, and only proper pairs (-f2) were selected.  A spike-in (RNA) normalization strategy was used essentially as described (Schwalb et al., 2016). This spike-in normalization allows for calculating sequencing depth σ_j, cross-contamination rate ϵ_j, and antisense bias ratio c_j. Sequencing depth σ_j allows to observe global shifts. The read counts (kij) for spike-ins were calculated using HTSeq (Anders et al., 2015).
mNET-seq: For all samples, paired-end 45 base reads were trimmed for adapters (-a TGGAATTCTCGGGTGCCAAGG -A GATCGTCGGACT), and low-quality bases (< Q20) were removed with Cutadapt (v 1.9.1) (Martin, 2011). The reads were then mapped to the GRCh38.p13 genome assembly merged with the yeast genome version SacCer3 using STAR (v 2.5.2b) (Dobin et al., 2013) with a maximum of 2% mismatches [mapping parameters: --outFilterMultimapScoreRange 0 --outFilterMismatchNoverLmax 0.02 --outFilterMatchNmin 16 --outFilterScoreMinOverLread 0  --outFilterMatchNminOverLread 0 --alignIntronMax 500000 ]. Samtools (v 1.3.1) (Li et al., 2009) was used to quality filter SAM files, where alignments with MAPQ smaller than 7 (-q 7) were skipped, and only proper pairs (-f 2) were selected.   Read counts (kij) for yeast genes for each condition j (control or CDK7as inhibited) were calculated using HTSeq (Anders et al., 2015). The counts data was used to generate normalization factors with DESeq2 (Love et al., 2014) to normalize the mNET-seq data.
MNase-seq: For all samples, paired-end 45 base reads were trimmed for low-quality bases, random matches, and for a minimum length [trimming parameters: -q 20,20 -O 12 -m 25] with Cutadapt (v 1.9.1) (Martin, 2011). Processed reads were then mapped to the GRCh38.p13 genome assembly using STAR (v 2.6.1b) (Dobin et al., 2013) with maximum 2 percent mismatches [mapping parameters: --outFilterMismatchNoverLmax 0.02 --outFilterMultimapScoreRange 0 --alignIntronMax 1]. Samtools (v 1.3.1) (Li et al., 2009) was used to quality filter SAM files, where alignments with MAPQ smaller than 7 (-q 7) were skipped, and only proper pairs (-f 2) were selected.
MNase-ChIP-seq: For all samples, paired-end 43 base reads were mapped using Bowtie 2 (v 2.3.4.1) (Langmead and Salzberg, 2012) to the merged human hg38 (GRCh38.p13) and Drosophila (BDGP6.28) genome assembly [mapping parameters: --no-discordant --no-mixed --very-sensitive]. SAMtools (v 1.3.1) (Li et al., 2009) was used to quality filter SAM files, whereby alignments with MAPQ smaller than 7 (-q 7) were skipped, and only proper pairs (-f2) were selected. Further data processing was carried out using the R/Bioconductor environment. Duplicate fragments and fragments longer than 100 bp (70 bp for Pol II) were excluded from further analysis. MNase-ChIP-seq coverages were obtained from piled-up fragment midpoint counts for every genomic position. Pol II, Ser5P, and SPT5 coverages were normalized to the signal at untranscribed regions to allow for the detection of global changes. To this end, genome-wide coverage was calculated from all MNase-ChIP-seq fragment midpoints in consecutive 2 kbp bins throughout the non-transcribed human genome (i.e. excluding 32,789 transcribed units we annotated using GenoSTAN (Zacher et al., 2017) and everything annotated in GENCODE v31) and used to generate normalization factors with DESeq2 (Love et al., 2014). For all other factors, which exhibited a peak-like binding behavior, normalization factors were obtained from total human fragment counts. Bigwig files are for normalized (except Inputs), entire fragment coverages.
Assembly: GRCh38
Supplementary files format and content: bigwig files

molecule: genomic DNA

organism: Homo sapiens

source_name: genome edited HEK293

growth_protocol: Cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM, Gibco, 11960085) supplemented with 10% fetal bovine serum (Gibco, 10500–064) and 1 % GlutaMAX (Thermo Fisher Scientific, 35050087) in a humidified incubator at 37°C and 5% CO2. Cells were passaged every 2-3 days using 0.25% Trypsin-EDTA (Gibco, 25200056). Biological replicates were cultured separately. All cell lines were verified to be free of mycoplasma contamination with PlasmoTest Mycoplasma Detection Kit (InvivoGen).   Drosophila Schneider-2 (S2) cells, used for MNase-ChIP-seq spike-ins, were obtained from DSMZ (ATCC Cat# CRL-1963, RRID: CVCL_Z232) and cultured in Schneider's Drosophila-Medium (Gibco, 21720024) supplemented with 10% fetal bovine serum (Gibco, 10500–064) in a non-humidified incubator at 27°C, protected from light.  Yeast S. cerevisiae cells (BY4741 strain), used for mNET-seq spike-ins, were obtained from Euroscarf (ACC-Y00000) and cultured in yeast medium (YP + 2% glucose) at 30°C until OD595 of 0.8 was reached.

extract_protocol:
TT-seq: Total RNA was extracted using TRIzol reagent (Invitrogen, 15596026), 4sU-labeled RNA was extracted according to Gressel at al., protocols.io, 2019.
mNET-seq: mNET-seq was performed as previously described (Caizzi et al. 2021; Gressel et al. 2017; Nojima et al. 2016; Schlackow et al. 2017) with minor changes. 50 mio CDK7as cells were treated for 15 or 30 min with 7.5 μM of 1-NM-PP1 or DMSO (solvent control). After the treatment, chromatin isolation was performed as described in the protocol for cellular fractionation (Conrad et al. 2017) using 25 mio cells per reaction. All buffers were supplemented with 7.5 μM 1-NM-PP1 or respective amounts of DMSO, protease (Sigma-Aldrich, P8340), and phosphatase (Roche, 4906837001) inhibitors. After the fractionation, the chromatin pellet was digested in 100 μl MNase digest buffer (1x MNase buffer, 1x BSA, 100 mM NaCl, 50 units/μl MNase (NEB, M0247S)) at 37°C and 1,400 rpm for 2 min. The digestion was stopped by adding 11 μl of 250 mM EGTA, and samples were centrifuged at 13,000 g for 5 min at 4°C. Supernatants were pooled together for each sample and 8-fold diluted with IP buffer (50 mM Tris HCl pH 7.4, 150 mM NaCl, 0.05% NP-40, 0.3% empigen (Sigma-Aldrich, 30326)). 15 μg of anti-Pol II CTD antibody (Diagenode, C15200004) conjugated with 125 μl of Dynabeads M-280 Sheep Anti-Mouse IgG (Invitrogen, 11201D) were added to each sample and incubated on a rotating wheel at 4°C and 12 rpm for 1 h. After IP, beads were washed seven times with 1 ml of IP buffer and once with 300 μl of PNKT buffer (1x T4 PNK buffer (NEB, M0236L), 0.1% Tween-20). After washes, 5’ RNA ends were phosphorylated with T4 Polynucleotide Kinase (PNK) (3' phosphatase minus; NEB, M0236L) at 800 rpm for 10 min at 37°C. Next, beads were washed once with IP buffer, and 0.5 ml of TRIzol reagent (Invitrogen, 15596026) was added to the beads. 2.5 ng of S. cerevisiae spike-ins were added to each sample in TRIzol. RNA spike-ins for mNET-seq were obtained from S. cerevisiae cells as previously described (Caizzi et al. 2021). RNA was extracted from beads, followed by RNA precipitation with 100% ethanol and GlycoBlue coprecipitant (Invitrogen, AM9515) at -80°C overnight. RNA pellets were washed twice with 70% ethanol, air-dried, and dissolved in 7 M urea. In order to size select 25-110 nt RNA, samples were resolved on 6% polyacrylamide gel containing 7 M urea. RNA of corresponding sizes was extracted from the gel and precipitated with 100% ethanol and GlycoBlue coprecipitant.
MNase-seq: 5 mio CDK7as cells were treated for 15 min with 7.5 μM of 1-NM-PP1 or DMSO (solvent control) and centrifuged at 500 g for 3 min at 4°C. Cell pellets were resuspended in 50 μl of Buffer I (10 mM Tris-HCl pH 7.5, 10 mM NaCl, 2.5 mM MgCl2, 0.5% NP-40, 10 % sucrose, 0.5 mM CaCl2) and incubated at 37°C for 2 min. Next, we added 500 μl of Buffer II (50 mM Tris-HCl pH 7.5, 50 mM NaCl, 2.5 mM MgCl2, 0.5% NP-40, 10 % sucrose, 2 mM CaCl2) supplemented with 3,000 U of MNase (NEB, M0247S) and incubated at 37°C for 9 min. MNase digestion was quenched with 11 μl of EGTA (500 mM). 550 μl of SDS Lysis Buffer (50 mM Tris-HCl pH 8.0, 1% SDS, 10 mM EDTA) was added, and samples were incubated on ice for 10 min. Samples were then treated with 0.1 μg/μl RNAse A (Thermo Scientific, EN0531) for 1.5 h at 37°C and 0.05 μg/μl Proteinase K (Life Technologies, AM2546) for 4 h at 65°C. DNA was extracted with an equal volume of phenol/chloroform/isoamyl alcohol (Sigma-Aldrich, P2069), followed by DNA precipitation with an equal volume of 100% ethanol and 200 mM NaCl for 2 h at -20°C. DNA pellets were washed twice with 1 ml of 70% ethanol, air-dried, and dissolved in water.
MNase-ChIP-seq: MNase-ChIP-seq was performed as previously described (Skene et al. 2015) with the following changes. 40 mio CDK7as cells were treated for 30 min with 7.5 μM of 1-NM-PP1 or DMSO (solvent control), following 8 min crosslinking with 1% formaldehyde (16% formaldehyde (w/v), methanol-free, Thermo Fisher Scientific, 28908). The crosslinking was quenched for 5 min with 125 mM glycine, and cells were washed with 10 ml PBS at room temperature. 2 ml of ice-cold PBS was added to plates for cell scraping. Scraped cells were transferred to falcons, washed twice with 5 ml ice-cold PBS, and centrifuged at 200 g for 5 min at 4°C. All buffers were supplemented with protease (Sigma-Aldrich, P8340) and phosphatase (Roche, 4906837001) inhibitors. Cell pellets were lysed with 5 ml of Farnham Lysis buffer (5 mM Pipes pH 8, 85 mM KCl, 0.5% NP-40) for 10 min on ice. Nuclei were pelleted by centrifugation at 1,700 g for 5 min at 4°C and washed with ice-cold PBS. Nuclei pellets were resuspended in 200 μl of 1% SDS Lysis buffer (50 mM Tris HCl pH 8.0, 10 mM EDTA, 1% SDS) and incubated for 10 min at room temperature to permeabilize nuclei. The reaction was quenched with 1.8 ml of IP buffer (20 mM Tris HCl pH 8.0, 150 mM NaCl, 1 mM EDTA, 1% Triton X-100) supplemented with 5 mM CaCl2 and incubated for 2 min at 37°C before the addition of 16,000U of MNase (NEB, M0247S) for 20 min at 37°C. MNase digestion was quenched with 40 μl of EDTA (500 mM) and 20 μl EGTA (500 mM). 1 ml of sample was transferred to AFA milliTubes (Covaris, 520135) and sonicated with a Covaris S220 Focused-ultrasonicator with the following parameters: 5% duty cycle, 140 W peak incident power, 200 cycles per burst, 600 sec processing time, 6°C bath temperature, continuous degassing mode, water level 8. Sonicated samples were centrifuged at 10,000 g for 15 min at 4°C, and supernatants were snap-frozen in liquid nitrogen and stored at −80°C before use. For each IP, 30 μg of sample chromatin and 200 ng of Drosophila S2 MNase-digested crosslinked chromatin were diluted in IP buffer to obtain a final concentration of 0.05% SDS and a final volume of 1.6 ml. Pre-clearing was carried out by adding 20 μl of Dynabeads Protein G (Thermo Fisher Scientific, 10009D) diluted in IP buffer, and incubating for 30 min at 4°C on the rotation wheel. Pre-cleared samples were incubated with 10 μg of Pol II (RPB1, Diagenode, C15200004), 10 μg of Ser5P (phospho-CTD Ser-5 of RPB1, Merck Millipore, 04-1572), 8 μl of Mediator (MED26, Cell Signaling, 14950), 6 μg of TFIIB (Santa Cruz, sc-271736), 5 μg of TFIIE (TFIIE-α, abcam, ab28177), 6 μg of NELF (NELF-E, Bethyl Laboratories, A301-913A) or 6 μg of SPT5 (Proteintech, 16511-1-AP) together with 1 μg of Spike-in (Active Motif, 61686) antibodies at 4°C overnight. The next day samples were centrifuged at 16,000g for 2 min at 4°C and inputs were taken for each sample. Samples were incubated for 2 h at 4°C with Dynabeads Protein G. Beads were washed once with Buffer 1 (20 mM Tris HCl pH 8.0, 150 mM NaCl, 2 mM EDTA, 1% Triton X-100, 0.1% SDS), four times with Buffer 2 (20 mM Tris HCl pH 8.0, 500 mM NaCl, 2 mM EDTA, 1% Triton X-100, 0.1% SDS), once with Buffer 3 (10 mM Tris HCl pH 8.0, 250 mM LiCl, 1 mM EDTA, 1% NP-40, 1% sodium-deoxycholate), and three times with TE buffer (10 mM Tris HCl pH 8.0, 50 mM NaCl, 1 mM EDTA). After the last wash, 200 μl of elution buffer (0.1 M NaHCO3, 160 mM NaCl, 1% SDS) was added to the beads, and samples were de-crosslinked overnight at 65°C. Next, samples were diluted with 200 μl of PBS and treated with 0.02 μg/μl RNAse A (Thermo Scientific, EN0531) for 1.5 h at 37°C and 0.2 μg/μl Proteinase K (Life Technologies, AM2546) for 2 h at 45°C. DNA was extracted with an equal volume (400 μl) of phenol/chloroform/isoamyl alcohol (Sigma-Aldrich, P2069), followed by DNA precipitation with 100% ethanol and 200 mM NaCl for 1 h at -80°C. DNA pellets were washed twice with 70% ethanol, air-dried, and dissolved in water.
TT-seq: Libraries were prepared from 100 ng of RNA with the strand-specific Ovation Universal RNA-Seq library kit (NuGEN, 0343-32).
mNET-seq: 8.4 ng of RNA for replicate 1 and 12.3 ng of RNA for replicate 2 were used for library preparation with TruSeq Small RNA Library Prep Kit (RS-200-0048) according to the manufacturer’s instructions.
MNase-seq: Libraries were prepared with the NEBNext Ultra II DNA Library Prep Kit for Illumina (NEB, E7645S) according to the manufacturer’s instructions.
MNase-ChiP-seq: Libraries were prepared with the NEBNext Ultra II DNA Library Prep Kit for Illumina (NEB, E7645S) according to the manufacturer’s instructions with adjusted for short fragments end repair, 5’ phosphorylation, and dA-tailing thermocycler program (30 min at 20°C, 1 h at 50°C; the heated lit set to 60°C). After amplification, libraries were double-sided (1.0 – 1.6x) size-selected with Agencourt AMPure XP beads (Beckman Coulter, A63881).

treatment_protocol: Cells were treated for 15, 30, or 60 min with 7.5 μM of 1-NM-PP1 (Sigma-Aldrich, St. Louis, MO USA) or an equal DMSO (solvent control; Sigma-Aldrich, St. Louis, MO USA) volume.

tag: cell line

value: genome edited HEK293

tag: cell type

value: Human embryonic kidney

tag: genotype

value: analog-sensitive CDK7

tag: treatment

value: 30 min 1-NM-PP1

tag: immunoprecipitation antibody

tag: method

value: MNase-ChIP-seq


title: HEK293CDK7as_MNaseChIPseq_Input_MED26_TFIIE_TFIIB_NELF_1-NM-PP1_30min_Rep2


title: ACS-1021 iPS-derived Cardiomyocytes 2

library_source: transcriptomic

molecule: total RNA

tag: tissue


title: hypothalamus, 120 day, M4-2

city: Tai'an

first: Qing

last: Li

address: Tai’an, Shandong Province, P.R. China

country: China

institute: College of Animal Science and Veterinary Medicine

data_processing:
Clean data (clean reads) were obtained by trimming reads containing adapter, ploy-N or with low quality from raw data
Index of the reference genome was built using hisat2-v2.0.5  and paired-end clean reads were mapped to the reference genome.
Stringtie-1.3.3b was used to count the read numbers mapped to each gene.
Assembly: ARS1.2
Supplementary files format and content: tab-delimited text files include FPKM values for each Sample.

organism: Capra hircus

source_name: hypothalamus

extract_protocol:
RNA was harvested using Trizol reagent.
A total of 1 µg total RNA per sample was used as input material for the lncRNA library preparation. Strand-specific libraries were generated using NEBNext® UltraTM RNA Library Prep Kit for Illumina® (NEB, USA) following manufacturer’s recommendations and index codes were added to attribute sequences to each sample

value: hypothalamus


title: N2, PE, biol rep 1

city: Chapel Hill

first: Robert

middle: Houston

last: Dowen

state: NC

address: 321 Fordham Hall, CB7100

department: Cell Biology and Physiology

country: USA

institute: University of North Carolina at Chapel Hill

description:
Column names listed in line one for each txt file
FC_gene_read_counts_tir-1_study.txt
DESeq2_norm_gene_counts_tir-1_study.txt

data_processing:
The adapter-trimmed mRNA-Seq reads were mapped to the WS260 genome using the STAR aligner. The mapping was performed using a GTF obtained from Wormbase (c_elegans.PRJNA13758.WS260.canonical_geneset.gtf) with the parameter --outFilterIntronMotifs RemoveNoncanonical.
The gene-level read counts were calculated using the featureCounts algorithm of the subread package (Liao et al., Bioinformatics, 2014), using the following flags: -t exon -g gene_id -a c_elegans.PRJNA13758.WS260.canonical_geneset.gtf.
Differential expression was calculated relative to the N2 wild-type sample using the DESeq2 package using a 1% FDR cutoff (Love et al., 2014). Comparisons between individual mutants were also performed.
Assembly: WS260
Supplementary files format and content: FC_gene_read_counts_study_name.txt: A tab-delimited text file for each sub-study that includes gene level unnormalized read counts for each sample in matrix format generated using the featureCounts aligorithm.
Supplementary files format and content: DESeq2_norm_gene_counts_study_name.txt: A tab-delimited text file that for each sub-study includes gene level normalized read counts for each sample in matrix format generated using the DESeq2 aligorithm.
Supplementary files format and content: DESeq2_DEGs_sample1_v_sample2_study_name.txt: A tab-delimited text file that includes a list of differentially expressed genes, the log2 FC values, and the p values called by DESeq2 at a 1% FDR threshold for sample1 relative to sample2 for each sub-study.

molecule: polyA RNA

organism: Caenorhabditis elegans

source_name: Whole animals

growth_protocol: Wild-type and mutant C. elegans animals were synchronized at the L1 stage by egg prep and grown for ~72 hours at 20°C on 10cm NGM Agarose plates (5000 animals/plate) seeded with E. coli OP50 until they reached day 1 of adulthood. For RNAi experiments, synchronized animals were reared on E. coli HT115 bacteria expressing dsRNAs for ~72 hours at 20°C on 10cm NGM Agarose plates containing 5 mM isopropyl-β-D-thiogalactoside (IPTG) and 100 μg/ml ampicillin.

extract_protocol:
Animals were harvested in M9 buffer, washed three times, and flash frozen in liquid Nitrogen. Total RNA was isolated using Trizol.
mRNA-Seq libraries were prepared using the Illumina TruSeq RNA Sample Preparation v2 kit. For the tra-1 and tir-1 studies, mRNA-Seq libraries were prepared by Novogene Inc.

tag: strain

value: wild-type

value: Standard growth conditions at 20C on NGM plates containing E. coli OP50

tag: developmental stage

value: Day 1 adults

value: Whole animals

tag: Sex

value: hermaphrodite


title: Mou_HeaC_R3

city: 昆明市

first: Lin

last: dongdong

state: 云南

address: 云南省昆明市五华区茭菱路

institute: Peking Union Medical College

data_processing:
Raw sequencing data was first filtered by Trimmomatic (version 0.36), low-quality reads were discarded and the reads contaminated with adaptor sequences were trimmed. Clean Reads were further treated with in-house scripts to eliminate duplication bias introduced in library preparation and sequencing. In brief, clean reads were first clustered according to the UMI sequences, in which reads with the same UMI sequence were grouped into the same cluster. Reads in the same cluster were compared to each other by pairwise alignment, and then reads with sequence identity over 95% were extracted to a new sub-cluster. After all sub-clusters were generated, multiple sequence alignment was performed to get one consensus sequence for each sub-clusters. After these steps, any errors and biases introduced by PCR amplification or sequencing were eliminated.
Deduplicated Reads were mapped to the reference genome of macaca mulatta and mus Musculus from Mmul_8.0.1 and GRCm38 respectively using STRA software (version 2.5.3a) with default parameters. Reads mapped to the exon regions of each gene were counted by featureCounts (Subread-1.5.1; Bioconductor) and then RPKM was calculated. Genes differentially expressed between groups were identified using the edgeR package (version 3.12.1). p-value cutoff of 0.05 and Fold-change cutoff of 2 were used to judge the statistical significance of gene expression differences. Gene ontology (GO) analysis and Kyoto encyclopedia of genes and genomes (KEGG) enrichment analysis for differentially expressed genes were both implemented by KOBAS software (version: 2.1.1) with a P-value cutoff of 0.05 to judge statistically significant enrichment. Alternative splicing events were detected by using rMATS (version 3.2.5) with an FDR value cutoff of 0.05 and an absolute value of Δψ of 0.05.
For lncRNA analysis, new lncRNAs were predicted. briefly, The mapped reads were spliced to obtain new transcripts by Stringtie(version 1.3.2). Then，new transcripts were fileted by four principle:1) Filter out the transcripts less than 200bp in length; 2) Filter out the transcripts containing less than 2 exons; 3) Include the transcripts containing at least 3 reads; 4) Remove known transcripts from the genome annotation file. Then, CPC2(version beta)、CPAT(version 1.2.4)、CNCI(version 2) and Pfam(version 27.0)were used to predict the coding potential of the selected transcripts. The transcripts predicted to be non-coding in all four softwares or database were referred to them as novel lncRNA.
The expression of novel lncRNA and genome annotated known lncRNA were counted by featureCounts (Subread-1.5.1; Bioconductor) and then RPKM was calculated. LncRNA differentially expressed between groups were identified using the edgeR package (version 3.12.1). P-value cutoff of 0.05 and fold-change cutoff of 2 were used to judge the statistical significance of lncRNA expression differences.
Assembly: Mmul_8.0; GRCm38

organism: Mus musculus

source_name: heart

extract_protocol:
Total RNAs were extracted from （monkey and mouse’ organs including heart, liver, spleen, lung, kidney, brain) using TRIzol reagent(Invitrogen, cat. NO 15596026) following the methods by Chomczynski et al (DOI:10.1006/abio.1987.9999). DNA digestion was carried out after RNA extraction by DNaseI. RNA quality was determined by examining A260/A280 with NanodropTM OneCspectrophotometer (Thermo Fisher Scientific Inc). RNA Integrity was confirmed by 1.5% agarose gel electrophoresis. Qualified RNAs were finally quantified by Qubit3.0 with QubitTM RNA Broad Range Assay kit (Life Technologies，Q10210).
2 μg total RNAs were used for stranded RNA sequencing library preparation using Ribo-off rRNA Depletion Kit (Human/Mouse/Rat) (Catalog NO. MRZG12324, Illumina) and KC-DigitalTM Stranded mRNA Library Prep Kit for Illumina® (Catalog NO. DR08502, Wuhan Seqhealth Co., Ltd. China) following the manufacturer’s instruction. The kit eliminates duplication bias in PCR and sequencing steps, by using unique molecular identifier (UMI) of 8 random bases to label the pre-amplified cDNA molecules. The library products corresponding to 200-500 bps were enriched, quantified and finally sequenced on DNBSEQ-T7 sequencer (MGI Tech Co., Ltd. China) with PE150 model.

value: heart

value: CHIKV control

tag: time


title: hypothalamus, 60 day, M2-5


title: HEK293CDK7as_MNaseChIPseq_Input_MED26_TFIIE_TFIIB_NELF_1-NM-PP1_30min_Rep1


title: ACS-1021 iPS-derived Cardiomyocytes 1


title: Mou_HeaC_R2

address: 云南省昆明市五华区茭菱路


title: ACS-1021 Vascularized Cardiac Organoid 3

value: ACS-1021 + RCEC

value: Various Cardiac


title: hypothalamus, 120 day, M4-1


title: hypothalamus, 60 day, M2-4


title: hypothalamus, 120 day, M4-3


title: ACS-1021 Vascularized Cardiac Organoid 2

value: Cardiac


title: Mou_BraT_R3

address: 云南省昆明市五华区茭菱路

source_name: brain

value: brain

value: CHIKV infection


title: HEK293CDK7as_MNaseChIPseq_Input_MED26_TFIIE_TFIIB_NELF_DMSO_30min_Rep3

value: genome edited HEK293

value: Human embryonic kidney

value: 30 min DMSO


title: hypothalamus, 60 day, M2-3


title: Mou_BraT_R2

address: 云南省昆明市五华区茭菱路


title: HEK293CDK7as_MNaseChIPseq_Input_MED26_TFIIE_TFIIB_NELF_DMSO_30min_Rep2

value: analog-sensitive CDK7

value: MNase-ChIP-seq


title: AU6844

city: Bellaterra

first: Ot

last: Massafret

state: Barcelona

address: carrer de la Vall Moronta, s/n

department: Departament de Biologia Cel·lular, Fisiologia i Immunologia

country: Spain

institute: Universitat Autònoma de Barcelona

data_processing:
The libraries were sequenced on a NovaSeq 6000 (Illumina) in paired-end mode with a read length of 2x51bp, following the manufacturer’s protocol for dual indexing.
RNA-seq reads were mapped against the human reference genome (GRCh38) using the STAR software version 2.7.8a with ENCODE parameters.
Annotated genes were quantified with RSEM v1.3.0 using default parameters and the human GENCODE annotation version 38.
Assembly: GRCh38

source_name: hES10 2iF

growth_protocol: blastomere and blastocyst-derived hESC lines were grown on feeder cells in KO-DMEM + 20% KSR medium supplemented with 2 mM L-glutamine, 1x MEM-non-essential amino acids, 50 mM 2-mercaptoethanol , 1x ITS-X, 50 U/mL penicillin - 50 ug/mL streptomycin and 4 ng/uL bFGF. Naïve hESCs were cultured in the same medium but supplemented with 10 ng/uL bFGF, 3 mM CHIR99021 and 0.5mmM PD0325901

extract_protocol:
RNA extraction was preformed using the Maxwell SimplyRNA Tissue kit (Promega)
RNA-seq libraries were prepared with the KAPA Stranded mRNA-Seq Illumina Platform Kit (Roche) following the manufacturer´s recommendations.

value: hES10 2iF

tag: group

value: naive

value: XX

value: Embryonic stem cell


title: hypothalamus, 60 day, M2-2

value: hypothalamus


title: AU6843

value: blastomere

value: XY


title: Mou_BraC_R3

address: 云南省昆明市五华区茭菱路

value: CHIKV control


title: hypothalamus, 60 day, M2-1


title: HEK293CDK7as_MNaseChIPseq_Input_1-NM-PP1_30min_Rep2

value: 30 min 1-NM-PP1


title: Mou_BraC_R2

address: 云南省昆明市五华区茭菱路


title: hypothalamus, 1 day, D1-4


title: AU6842


title: Mou_HeaC_R1

address: 云南省昆明市五华区茭菱路

value: heart


title: Mou_BraC_R1

address: 云南省昆明市五华区茭菱路

value: brain


title: HEK293CDK7as_MNaseChIPseq_Input_DMSO_30min_Rep3


title: hypothalamus, 1 day, D1-3


title: AU6841


title: HEK293CDK7as_MNaseChIPseq_Input_1-NM-PP1_30min_Rep1


title: AU6840


title: AU6839

value: blastocyst


title: hypothalamus, 1 day, D1-2


title: hypothalamus, 1 day, D1-1


title: HEK293CDK7as_MNaseChIPseq_Input_DMSO_30min_Rep2

value: genome edited HEK293


title: ACS-1021 Vascularized Cardiac Organoid 1

value: Cardiac

value: ACS-1021 + RCEC

value: Various Cardiac


title: HEK293CDK7as_MNaseChIPseq_Input_MED26_TFIIE_TFIIB_NELF_DMSO_30min_Rep1

value: Human embryonic kidney

value: analog-sensitive CDK7

value: 30 min DMSO

value: MNase-ChIP-seq


title: HEK293CDK7as_MNaseChIPseq_Input_DMSO_30min_Rep1


title: Mou_BraT_R1

address: 云南省昆明市五华区茭菱路

value: CHIKV infection


title: Zbp1_Nec1s2

city: Wuhan

first: Jinyang

last: Gu

address: 1277 Jiefang avenue

institute: Union hospital, Tongji medical college, Huazhong university of science and technology

data_processing:
CLC Genomics Workbench v 11.0.1
Sequence reads were trimmed for adapter sequence/low quality sequence using CLC Genomics Workbench (parameter-Quality limit: 0.05).
Trimmed sequence reads were mapped to GRCm39/mm39 using CLC Genomics Workbench (parameters-mismatch cost: 2 insertion cost: 3 deletion cost: 3 length fraction: 0.8 similarity fraction: 0.8)
Read count extraction and normalization were performed using CLC genomic workbench.
Assembly: mm39
Supplementary files format and content: tab-delimited text files include raw counts for each sample.

source_name: Liver

extract_protocol:
RNA was extracted by Trizol method.
1) By utilizing the structural characteristic that most eukaryotic mRNAs carry a polyA tail, mRNAs with polyA tails were enriched by Oligo(dT) magnetic beads; 2) The purified mRNAs were cleaved into small fragments with fragmentation buffer at a suitable temperature; 3) First-strand cDNAs were produced by reverse transcription using a random hexamer primer; 4) Second-strand cDNAs are synthesized (strand-specific library: dUTPs were used instead of dTTPs in the second-strand synthesis to incorporate dUTPs in the second-strand cDNAs, while the high-fidelity DNA polymerase used in this method could not amplify uracil-containing DNA templates, thus realizing the strand specificity), while simultaneously performing end repair and dA-Tailing; 5) Sequencing adapter ligation was performed, followed by DNA magnetic bead purification and fragment selection after ligation was completed to yield a library with 250-350 bp insert fragments; 6) The ligated products were amplified by PCR and purified again using DNA magnetic beads, where the products were solubilized with nuclear-free water; 7) After the initial library was constructed, a Qubit fluorescence quantifier was used for concentration detection, followed by a Qsep400 high-throughput biofragment analyzer for fragment size detection; 8) The double-stranded DNA library obtained by PCR was subjected to single-stranded cyclization to obtain a single-stranded circular DNA library; 9) The final library was amplified with phi29 before uploading to make DNA nanoballs (DNB) with more than 300 copies of a molecule, and the DNBs were loaded into the sequencing chip and sequenced on the BGI sequencing platform.

value: Liver

value: WT

value: ND; AAV8-Zbp1; Nec-1s; I/R


title: Zbp1_Nec1s1


title: HEK293CDK7as_MNaseChIPseq_Input_1-NM-PP1_30min_Rep3


title: hypothalamus, 1 day, D1-5

value: hypothalamus


title: AU6838

value: XY

value: Embryonic stem cell


title: HEK293CDK7as_MNaseChIPseq_SPT5_1-NM-PP1_30min_Rep3

value: 30 min 1-NM-PP1

value: SPT5 (Proteintech, 16511-1-AP)


title: AU6837

value: blastocyst


title: Yki, wing disc tumors, rep2

city: Hong Kong

first: Yan

last: Yan

address: Clear Water Bay

department: LIFS

institute: HKUST

description: 10x Genomics

data_processing:
Raw data mapping and primary analysis was performed in the Cell Ranger pipeline.
Secondary analysis for marker gene expression pattern was performed with Seurat (Butler et al., 2018).
Assembly: GCF_000001215.4
Supplementary files format and content: csv files include normalized abundance measurements for each Sample

organism: Drosophila melanogaster

source_name: Wing imaginal disc

growth_protocol: To generate MARCM clones for each genotype, around 50 embryos were collected within 3 hours and raised until 60 hours after egg laying (AEL). The larvae were then placed in water bath for 1 hour at 37°C for heat shock and dissected at 36 hours to perform single cell experiment.

extract_protocol:
The wing imaginal discs were dissociated in 0.25% Trypsin-EDTA solution at 37 ℃ for 10 min. Cells were then washed in DPBS and passed through 35μm filter before library preparation.
Construction of 10x single cell libraries and sequencing on Illumina Hiseq platform were performed by Novogene.

value: Wing imaginal disc cells

value: Wing imaginal disc

value: hsFlp;Tub>y+>Gal4 UAS-GFP/UAS-YkiS168A; Tub-Gal80 FRT82B/scrib1FRT82B

tag: age

value: 96 hours after egg laying


title: Vector_Nec1s3

value: ND; AAV8-Vector; Nec-1s; I/R


title: HEK293CDK7as_MNaseChIPseq_SPT5_1-NM-PP1_30min_Rep2

value: genome edited HEK293

value: Human embryonic kidney

value: analog-sensitive CDK7


title: Zbp1_Nec1s3

value: Liver


title: RasV12_2nd, wing disc tumors, rep2

description: 10x Genomics

value: hsFlp;Tub>y+>Gal4 UAS-GFP/UAS-RasV12; Tub-Gal80 FRT82B/scrib1FRT82B


title: RasV12_1st, wing disc tumors, rep1

description: 10x Genomics


title: AU6836

value: XX


title: HEK293CDK7as_MNaseChIPseq_SPT5_DMSO_30min_Rep3

value: 30 min DMSO

value: MNase-ChIP-seq


title: Vector_Nec1s1

value: WT


title: HepG2 cells, RbIgG-RIP, rep2

city: Jinan

first: Jinyuan

last: Liang

address: wenhua road

institute: Shandong university

description: PABPC4 versus IgG.txt

data_processing:
In silico ribosomal depletion was performed by mapping the reads to human rRNA (National Center for Biotechnology Information) using bowtie2 (version 2.4.1).
The remaining reads were aligned to human reference genome GRCh38 using STAR (version 2.7.10a).
After removed the PCR duplication, quantification were analyzed by featureCounts (version 2.0.1).
Assembly: hg38
Supplementary files format and content: The differential binding sites were provided as txt data files.

growth_protocol: HepG2 cells were maintained in DMEM supplemented with 15% fetal bovine serum (FBS) and antibiotics in humidified atmosphere with 5% CO2 at 37 °C.

extract_protocol:
HepG2 cell pellet was lysed and RNA was immunoprecipitated with pre-prepared antibody conjugated protein A/G magnetic beads. After washing, RNA was extracted by TRI Reagent and used for further sequencing.
Libraries were prepared using EpiTM mini longRNA-seq kit following manufacturer's protocols.
SMART-seq use random primers

value: hepatocellular carcinoma cells

tag: rip antibody

value: anti-IgG (Sigma-Aldrich, PP64)

value: PABPC4 overexpressed


title: HEK293CDK7as_MNaseChIPseq_SPT5_1-NM-PP1_30min_Rep1

value: 30 min 1-NM-PP1

value: SPT5 (Proteintech, 16511-1-AP)


title: NICD_1st, wing disc tumors, rep1

description: 10x Genomics

value: Wing imaginal disc cells

value: Wing imaginal disc

value: hsFlp; Tub>y+>Gal4 UAS-GFP/UAS-NICD; Tub-Gal80 FRT82B/scrib1FRT82B


title: HepG2 cells, RbIgG-RIP, rep1

description: PABPC1 versus IgG.txt

value: PABPC1 overexpressed


title: NICD_2nd, wing disc tumors, rep2

description: 10x Genomics

value: 96 hours after egg laying


title: HepG2 cells, PABPC4-RIP, rep2

description: PABPC4 versus IgG.txt

value: PABPC4 (Proteintech, 14960-1-AP)


title: Zbp1_Vehicle2

value: Liver

value: ND; AAV8-Zbp1; Vehicle; I/R


title: HEK293CDK7as_MNaseChIPseq_SPT5_DMSO_30min_Rep2

value: genome edited HEK293

value: Human embryonic kidney

value: analog-sensitive CDK7


title: HEK293CDK7as_MNaseChIPseq_SPT5_DMSO_30min_Rep1

value: 30 min DMSO

value: MNase-ChIP-seq


title: Zbp1_Vehicle1

value: WT


title: Mscrib_1st, wing disc tumors, rep1

description: 10x Genomics

value: hsFlp;Tub>y+>Gal4 UAS-GFP/+; Tub-Gal80 FRT82B/scrib1FRT82B


title: HepG2 cells, PABPC1-RIP, rep2

description: PABPC1 versus IgG.txt

value: hepatocellular carcinoma cells

value: PABPC1 (Proteintech, 10970-1-AP)


title: M82B, wild type wing discs

description: 10x Genomics

value: Wing imaginal disc cells

value: hsFlp;Tub>y+>Gal4 UAS-GFP/+; Tub-Gal80 FRT82B/FRT82B (FRT82B #2035 from BDSC)


title: HEK293CDK7as_MNaseChIPseq_NELF-E_1-NM-PP1_30min_Rep3

value: 30 min 1-NM-PP1

value: NELF-E (Bethyl Laboratories, A301-913A)


title: Vector_Vehicle3

value: ND; AAV8-Vector; Vehicle; I/R


title: Mscrib_2nd, wing disc tumors, rep2

description: 10x Genomics

value: Wing imaginal disc

value: 96 hours after egg laying


title: Vector_Nec1s2

value: Liver

value: ND; AAV8-Vector; Nec-1s; I/R


title: HEK293CDK7as_MNaseChIPseq_NELF-E_1-NM-PP1_30min_Rep1

value: genome edited HEK293

value: Human embryonic kidney


title: HepG2 cells, sgPABPC4_2, rep2

data_processing:
The adaptor sequences and low-quality sequence reads were removed from the data sets. Raw sequences were  transformed into clean reads after data processing. These clean reads were then mapped to the reference genome  sequence. Only reads with a perfect match or one mismatch were further analyzed and annotated based on the  reference genome. Hisat2 tools soft were used to map with reference genome.
The StringTie Reference Annotation Based Transcript (RABT) assembly method was used to construct and  identify both known and novel transcripts from Hisat2 alignment results
Assembly: hg38
Supplementary files format and content: FPKM and TPM values were provided in a tabular format.

source_name: HepG2 PABPC4-KO clone 2

extract_protocol:
Total RNA was extracted from the samples using the TRlzol Reagent (Lifetechnologies). 1.3 ug of total RNA was used for the construction of sequencing libraries.
RNA libraries for RNA-seq were prepared using Hieff NGS Ultima Dual-mode mRNA Library  Prep Kit for Illumina (Yeasen Biotechnology (Shanghai) Co., Ltd.) following manufacturer's protocols.

value: PABPC4 knockout


title: Vector_Vehicle1


title: Experiment_3_TYRO3_dADAM

city: Turku

first: Katri

middle: Mataleena

last: Vaparanta

address: Vanha Hämeentie 82 as 2

department: Institute of Biomedicine

country: Finland

institute: University of Turku

data_processing:
Base-calling with bcl2fastq2 conversion software (HiSeq).
Quality check with FastQC  v. 0.11.9
Reads trimmed with Trimmomatic v. 0.33 (parameters: TruSeq3-PE-2 2:30:10 ) and PRINSEQ v. 0.20.4 (parameters: left 13, right 3)
Pseudoalignment with kallisto v. 0.46.0, default parameters
Genome_build: GRCh38
Supplementary_files_format_and_content: tsv files containg raw counts and TPM (transcript per million) values

growth_protocol: The WM-266-4 cells were cultured in DMEM medium (Life Technologies) supplemented with 10% (weight/vol) fetal calf serum (FCS) (Promocell) and 1 μg/ml puromycin (Sigma-Aldrich).

extract_protocol:
Total RNA extraction with NucleoSpin RNA Plus kit (Macherey-Nagel) according to manufacturer's protocol.
The sequencing libraries were constructed with TruSeq Stranded mRNA HT Kit (Illumina) according to TruSeq® Stranded mRNA Sample Preparation, Illumina (15031047) protocol. IDT for Illumina TruSeq RNA UD Indices (Illumina) were used fo indexing.

treatment_protocol: The cells were transfected with jetPRIME (Polyplus-transfection) and serum starved overnight before RNA extraction.

value: Stable lentiviral TYRO3 knock-down

tag: transfection

value: pEZY-myc-his TYRO3 ΔADAM (P408A/L409P/G419P)

value: Melanoma cell line


title: Zbp1_Vehicle3

value: WT

value: ND; AAV8-Zbp1; Vehicle; I/R


title: Experiment_3_TYRO3_dGS

value: pEZY-myc-his TYRO3 ΔGS (I449A)


title: HEK293CDK7as_MNaseChIPseq_NELF-E_DMSO_30min_Rep3

value: analog-sensitive CDK7

value: 30 min DMSO

value: MNase-ChIP-seq


title: HepG2 cells, sgPABPC4_2, rep1

value: hepatocellular carcinoma cells


title: Experiment_3_TYRO3_WT

value: pEZY-myc-his TYRO3 wild-type


title: HEK293CDK7as_MNaseChIPseq_NELF-E_DMSO_30min_Rep2

value: NELF-E (Bethyl Laboratories, A301-913A)


title: HepG2 cells, PABPC4-RIP, rep1

description: PABPC4 versus IgG.txt

value: PABPC4 (Proteintech, 14960-1-AP)

value: PABPC4 overexpressed


title: Experiment_3_GFP

value: pEZY-myc-his GFP


title: HepG2 cells, sgPABPC4_1, rep2

source_name: HepG2 PABPC4-KO clone 1


title: HEK293CDK7as_MNaseChIPseq_NELF-E_DMSO_30min_Rep1

value: genome edited HEK293

value: Human embryonic kidney


title: HepG2 cells, sgPABPC4_1, rep1

value: PABPC4 knockout


title: Zbp1_KO1

value: Liver

value: HFD; I/R


title: HEK293CDK7as_MNaseChIPseq_NELF-E_1-NM-PP1_30min_Rep2

value: 30 min 1-NM-PP1


title: HepG2 cells, PABPC1-RIP, rep1

description: PABPC1 versus IgG.txt

value: PABPC1 (Proteintech, 10970-1-AP)

value: PABPC1 overexpressed


title: Zbp1_WT3


title: Experiment_2_TYRO3_dADAM

value: Stable lentiviral TYRO3 knock-down

value: pEZY-myc-his TYRO3 ΔADAM (P408A/L409P/G419P)

value: Melanoma cell line


title: HEK293CDK7as_MNaseChIPseq_TFIIE_1-NM-PP1_30min_Rep3

value: analog-sensitive CDK7

value: TFIIE-α (abcam, ab28177)

value: MNase-ChIP-seq


title: Experiment_2_TYRO3_dGS

value: pEZY-myc-his TYRO3 ΔGS (I449A)


title: Vector_Vehicle2

value: WT

value: ND; AAV8-Vector; Vehicle; I/R


title: HepG2 cells, sgRen, rep4

source_name: HepG2 sgRen control

value: hepatocellular carcinoma cells


title: HepG2 cells, sgRen, rep3


title: Zbp1_WT2


title: Experiment_2_TYRO3_WT

value: pEZY-myc-his TYRO3 wild-type


title: Zbp1_KO3


title: HepG2 cells, sgRen, rep2


title: HEK293CDK7as_MNaseChIPseq_TFIIE_1-NM-PP1_30min_Rep1

value: genome edited HEK293

value: Human embryonic kidney


title: HepG2 cells, sgRen, rep1


title: Experiment_1_TYRO3_dADAM


title: Zbp1_WT1

value: Liver

value: HFD; I/R


title: Experiment_2_GFP

value: pEZY-myc-his GFP


title: HEK293CDK7as_MNaseChIPseq_TFIIE_DMSO_30min_Rep3

value: 30 min DMSO


title: C4

city: WuHan

first: Yi-Xuan

last: Tu

state: HuBei

address: 1 Shizishan Rd

department: College of Life Science and Technology

institute: Huazhong agricultural university

data_processing:
FastQC was used to check the quality of the raw sequence fastq files. High-quality pruning was performed using fastp.
Paired end reads were mapped to the reference genome of Ensembl (GRCm39) using HISAT2. And SAMtools is used to sort and index aligned BAM files.
Reads were counted using the featureCounts  program in the Subread.
Assembly: GRCm39
Supplementary files format and content: tab-delimited text files show count values for each sample

source_name: intestinal tract

extract_protocol:
Following manufacturer's protocol, the total RNA of sample was extracted using Trizol reagent.
After Sample QC, the mRNA was hybridized with Oligo（dT）probe and Captured by Magnetic Beads. The mRNA was then reverse-transcripted to first strand DNA.First strand DNA was used as template to synthesis second strand DNA, obtaining dsDNA. The dTTP tailed adaptor was ligated to both ends of the dsDNA fragments. The ligation product was amplified by PCR, and circularized to get single-stranded circular(ssCir) library. The ssCir library was then amplified through rolling circle amplification (RCA) to obtain DNA nanoball(DNB). The DNB was then loaded to flowcell, and sequenced by DNBSEQ Platform.

value: intestinal tract

value: control


title: HEK293CDK7as_MNaseChIPseq_TFIIE_DMSO_30min_Rep2


title: Zbp1_KO2


title: C3


title: virus

city: Beijing

first: Yang

last: Jiang

state: Beijing

address: No.5, Dongdan Three Street, Dongcheng District, Beijing

department: Department of Immunology, Institute of Basic Medical Sciences

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

data_processing:
First, the original data were filtered and quality were controlled to ensure the accuracy of the sequencing data.
raw2clean_QC（v1.0）
Then, according to primer sequences provided by the ZhangFeng, the sequencing reads were screened and compared with sgRNA sequences in sgRNA library.
mageck（v0.5.9.3）
The next, the number of sgRNA and  its relative gene in each sample was counted, and after standardization, sample quality control were performed (sgRNA read counts distribution, boxplot, PCA, correlation heat map). Finally, differences of sgRNA between groups were compared and the relative genes were screened, and GSEA was conducted.
Assembly: hg18
Supplementary files format and content: tab-delimited text files include the  values of sgRNA for each Sample

source_name: cell line

growth_protocol: human amnionic WISH cells were maintained in DMEM supplemented with 10% fetal bovine serum (FBS) in humidified atmosphere with 5% CO2 at 37C

extract_protocol:
The genomic DNA was extracted by  TIANamp Genomic DNA Kit (DP304, Tiangen). all DNA was used toPCR for enrichment of sgRNA. The sequences of primers for sgRNA amplification were as follows: forward, 5’-AGAGATAATTAGAATTAATTTGACTGTAAACACAAAGATATTAG-3’; reverse, 5’-AAGTTGATAACGGACTAGCCTTATTTTAACTTGCTATTTCT-3’.
Firstly， the PCR products’  terminal were repaired and the 3’  A tail were added to the end, the DNA template was connected to the splice at both ends of the fragment, to prepare PCR-free library
The Human CRISPR Knockout Pooled Library sequnence

treatment_protocol: The Human CRISPR Knockout Pooled Library (GeCKO v2, 1000000048, Addgene) was transferred into WISH cells.After puromycin selection for 10 days,half of  these cells were infected with HSV-1 (MOI:10)  for 30 hours, and  the remaining lived infected cells and uninfected cells  were collected for sgRNA sequencing and enrichment analysis.

value: cell line

value: WISH

value: human amnionic WISH cells

value: HSV-1 infection for 30h


title: Experiment_1_TYRO3_dGS

value: Stable lentiviral TYRO3 knock-down

value: pEZY-myc-his TYRO3 ΔGS (I449A)

value: Melanoma cell line


title: C2


title: NC

value: WT

value: HSV-1 infection for 0h


title: Experiment_1_TYRO3_WT

value: pEZY-myc-his TYRO3 wild-type


title: HEK293CDK7as_MNaseChIPseq_TFIIE_1-NM-PP1_30min_Rep2

value: genome edited HEK293

value: Human embryonic kidney

value: analog-sensitive CDK7

value: 30 min 1-NM-PP1

value: TFIIE-α (abcam, ab28177)

value: MNase-ChIP-seq


title: HEK293CDK7as_MNaseChIPseq_TFIIB_1-NM-PP1_30min_Rep3

value: TFIIB (Santa Cruz, sc-271736)


title: mRNAs in CD133(positive) Osteosarcoma stem cells 6

city: beijing

first: Xiong

last: Shu

address: No.31, Xinjiekou East Street, Xicheng District, Beijing

institute: Beijing Research Institute of Traumatology and Orthopaedics, Beijing JiShuiTan Hospital

description: paired-end

data_processing:
The data volume was about 10 G/ sample, and the proportion of each base mass greater than 20 (Q20) was not less than 90%.
Hisat2 software is used in sequence alignment, and its alignment algorithm is called Spliced Maping.
Trimmed fragments were counted from each of the subsequent gene segments using Stringtie comparison, followed by normalization with TMM (TMM mean of M Values) algorithm, and then the FPKM value for each gene.
Assembly: GRCh38
Supplementary files format and content: tab-delimited text files include read count and FPKM values for each Sample

source_name: Saos-2 cells

extract_protocol:
In this project, miRNeasy Mini Kit Qiagen#217004 Kit was used for RNA extraction of samples. The extraction was carried out strictly in accordance with the standard operating procedure manual provided by the Kit manufacturer, and the total RNA of samples was extracted.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: Saos-2 cells

value: CD133(+) Osteosarcoma stem cells


title: C1

value: intestinal tract

value: control


title: B4

value: black rice


title: Experiment_1_GFP

value: Stable lentiviral TYRO3 knock-down

value: pEZY-myc-his GFP


title: mRNAs in CD133(negative) Osteosarcoma stem cells 6

description: paired-end

value: CD133(-) Osteosarcoma stem cells


title: HEK293CDK7as_MNaseChIPseq_TFIIB_1-NM-PP1_30min_Rep1


title: mRNAs in CD133(positive) Osteosarcoma stem cells 5

description: paired-end


title: HEK293CDK7as_MNaseChIPseq_TFIIB_1-NM-PP1_30min_Rep2


title: B3


title: mRNAs in CD133(negative) Osteosarcoma stem cells 5

description: paired-end


title: HME2_ECM_2

city: Baltimore

first: Daniele

middle: Marie

last: Gilkes

state: MD

address: 1650 N Orleans St, CRB1 128, None

department: Oncology

institute: Johns Hopkins

data_processing:
Preliminary analysis was performed by Quick Biology. The reads were then mapped to the latest UCSC transcript set using Bowtie2 version 2.1.0.
Gene expression level was estimated using RSEM v1.2.15.
TMM (Trimmed Mean of M-values) was used to normalize gene expression.
Differentially expressed genes were identified using the edgeR program.
Supplementary_files_format_and_content: Comma delimited text file containing gene-level counts

source_name: HME2_ECM from BJ fibroblasts

growth_protocol: MCF10A, PCS600010, htertHME, HME2 cells, and BJ fibroblasts were obtained from ATCC and cultured as recommended.

extract_protocol:
Total RNA was extracted from cells using TRIzol (Invitrogen) and purified using Direct-zol RNA mini kit (Zymo Research) with DNase I treatment.
Libraries for RNA-Seq were prepared with KAPA Stranded RNA-Seq Kit. The workflow consisted of mRNA enrichment, cDNA generation, end repair to generate blunt ends, A-tailing, adaptor ligation and 12 cycles of PCR amplification. Unique adaptors were used for each sample in order to multiplex samples into several lanes. Sequencing was performed on Illumina Hiseq 3000/4000 with a 150bp pair-end run. A data quality check was done on Illumina SAV. Demultiplexing was performed with Illumina Bcl2fastq2 v 2.17 program.

treatment_protocol: We cultured MCF10A, PCS600010, htertHME, and HME2 cells in decellularized ECM scaffolds generated by BJ fibroblasts or on an uncoated, plastic cell culture dish (Unc) to analyze the effects of ECM signalling on breast cancer development.

tag: culture method

value: ECM from BJ fibroblasts


title: HME2_ECM_1


title: HEK293CDK7as_MNaseChIPseq_TFIIB_DMSO_30min_Rep3

value: 30 min DMSO


title: B1


title: B2


title: mRNAs in CD133(positive) Osteosarcoma stem cells 4

description: paired-end


title: HME2_Unc_2

value: Uncoated


title: mRNAs in CD133(negative) Osteosarcoma stem cells 4

description: paired-end


title: C_M_AOM_3


title: HEK293CDK7as_MNaseChIPseq_TFIIE_DMSO_30min_Rep1

value: genome edited HEK293

value: Human embryonic kidney


title: C_M_AOM_2


title: HEK293CDK7as_MNaseChIPseq_MED26_1-NM-PP1_30min_Rep3

value: analog-sensitive CDK7

value: 30 min 1-NM-PP1

value: MED26 (Cell Signaling, 14950)


title: HME2_Unc_1


title: HEK293CDK7as_MNaseChIPseq_TFIIB_DMSO_30min_Rep2

value: TFIIB (Santa Cruz, sc-271736)

value: MNase-ChIP-seq


title: HME2_ECM_3


title: HEK293CDK7as_MNaseChIPseq_MED26_1-NM-PP1_30min_Rep2


title: lncRNAs in CD133(positive) Osteosarcoma stem cells 3

description: paired-end

value: Saos-2 cells

value: CD133(+) Osteosarcoma stem cells


title: C_M_AOM_1

value: intestinal tract

value: control


title: lncRNAs in CD133(negative) Osteosarcoma stem cells 3

description: paired-end


title: htertHME_ECM_1

source_name: htertHME_ECM from BJ fibroblasts

value: htertHME


title: HEK293CDK7as_MNaseChIPseq_MED26_1-NM-PP1_30min_Rep1


title: C_F_AOM_3


title: lncRNAs in CD133(positive) Osteosarcoma stem cells 2

description: paired-end


title: htertHME_ECM_3


title: htertHME_Unc_3


title: HEK293CDK7as_MNaseChIPseq_MED26_DMSO_30min_Rep3


title: lncRNAs in CD133(negative) Osteosarcoma stem cells 2

description: paired-end


title: C_F_AOM_2


title: HEK293CDK7as_MNaseChIPseq_TFIIB_DMSO_30min_Rep1


title: htertHME_Unc_2


title: lncRNAs in CD133(positive) Osteosarcoma stem cells 1

description: paired-end


title: C_F_AOM_1


title: htertHME_Unc_1


title: HEK293CDK7as_MNaseChIPseq_MED26_DMSO_30min_Rep2


title: PCS600010_ECM_3

source_name: PCS600010_ECM from BJ fibroblasts


title: lncRNAs in CD133(negative) Osteosarcoma stem cells 1

description: paired-end


title: HEK293CDK7as_MNaseChIPseq_MED26_DMSO_30min_Rep1

source_name: genome edited HEK293


title: HEK293CDK7as_MNaseChIPseq_Ser5P_CTD_RPB1_1-NM-PP1_30min_Rep3

value: phospho-CTD Ser-5 of RPB1 (Merck Millipore, 04-1572)


title: B_M_AOM_3

value: black rice


title: B_M_AOM_2


title: HEK293, 45min 4sU run-on repliacte3

city: Guangzhou

first: Jie

last: Zhang

address: ShiLouGang Road 10

institute: Institute of Biological and Medical Engineering， Guangdong Academy of Science

library_source: other

data_processing:
library strategy: Flux-seq
The data was first trimmed by fastp with option: -f 5 -U --umi_loc=read1 --umi_len=8 --umi_prefix=UMI --adapter_sequence=CTGTCTCTTATACACATCT --adapter_sequence_r2=CTGTCTCTTATACACATCT ; then aligned to hg38 by bowtie2 with option: --very-sensitive --no-unal; the sam files were then transfer to bam files and sorted by samtools. Then the data were transfer to bedgraph file by: genomeCoverageBed -bga -ibam
Assembly: hg38
Supplementary files format and content: bedfiles refer to the polymerase flux of genes

molecule: other

growth_protocol: HEK293 cells were maintained in DMEM supplemented with 10% fetal bovine serum (FBS) with 5% CO2 at 37C

extract_protocol:
RNA was harvested using miRNeasy Tissue/Cells Advanced Micro Kit (Qiagen)
RNA were fragmented, biotinlated, then enriched, ligated with adaptors, RT PCR, PCR

treatment_protocol: 1:200 of 200mM 4sU were added to each sample for 5min/15min/45min run-on

value: Human Embryonic Kidney Cells

value: 45min 4sU run-on


title: HEK293CDK7as_MNaseChIPseq_Ser5P_CTD_RPB1_1-NM-PP1_30min_Rep2


title: B_M_AOM_1


title: PCS600010_ECM_1

value: ECM from BJ fibroblasts


title: HEK293CDK7as_MNaseChIPseq_Ser5P_CTD_RPB1_1-NM-PP1_30min_Rep1

value: genome edited HEK293


title: PCS600010_ECM_2


title: HEK293, 45min 4sU run-on repliacte2


title: HEK293, 45min 4sU run-on repliacte1


title: B_F_AOM_3


title: PCS600010_Unc_3

value: Uncoated


title: HEK293CDK7as_MNaseChIPseq_Ser5P_CTD_RPB1_DMSO_30min_Rep3

value: Human embryonic kidney

value: analog-sensitive CDK7

value: 30 min DMSO


title: HEK293, 15min 4sU run-on repliacte3

value: 15min 4sU run-on


title: PCS600010_Unc_1


title: HEK293CDK7as_MNaseChIPseq_Ser5P_CTD_RPB1_DMSO_30min_Rep1

value: MNase-ChIP-seq


title: B_F_AOM_2


title: HEK293CDK7as_MNaseChIPseq_Ser5P_CTD_RPB1_DMSO_30min_Rep2


title: B_F_AOM_1


title: HEK293, 5min 4sU run-on repliacte3

value: 5min 4sU run-on


title: HEK293, 15min 4sU run-on repliacte2


title: HEK293CDK7as_MNaseChIPseq_PolII_RPB1_1-NM-PP1_30min_Rep2

value: 30 min 1-NM-PP1

value: Pol II CTD antibody (Diagenode, C15200004)


title: HEK293CDK7as_MNaseChIPseq_PolII_RPB1_1-NM-PP1_30min_Rep3


title: MCF10A_ECM_3

source_name: MCF10A_ECM from BJ fibroblasts


title: WT_M_Apc_2

value: intestinal tract


title: WT_M_Apc_3


title: HEK293, 5min 4sU run-on repliacte2


title: HEK293, 15min 4sU run-on repliacte1


title: HEK293, 5min 4sU run-on repliacte1


title: HEK293CDK7as_MNaseChIPseq_PolII_RPB1_1-NM-PP1_30min_Rep1


title: WT_F_Apc_3


title: HEK293CDK7as_MNaseChIPseq_PolII_RPB1_DMSO_30min_Rep3


title: PCS600010_Unc_2


title: MCF10A_Unc_3


title: WT_F_Apc_1


title: patient10

city: Chengdu

first: Wenchuan

last: Qi

state: Sichuan

address: 37,12th Bridge Road, Jinniu District

institute: Chengdu University of Traditional Chinese Medicine

description: exosomal microRNAs

data_processing:
1. Data quality control and filtering  The quality information of raw data in FASTQ format was calculated and then the raw data was filtered using the Personalbio company's self developed script, clean data was obtained by removing adapter and Low quality sequence. Filter Clean Reads from 18 nt to 36 nt in length and perform deduplicatio n to obtain Unique Reads for subsequent analysis.
2. Data mapping analysis  Reference genome index was built by Bowtie2 v 2.2.6) and the de duplicated Clean Reads were mapping to the reference genome using the miRDeep 2( v 2.0.0.8 )software.
3. Small RNA classification notes  Unique Reads is annotated with known miRNAs first in the miRBase database http://www.mirbase.org/ and then annotated with other non coding RNAs. Sequences that are not annotated with any information use mireap v 2.0 for new miRNA prediction analysis. Organize the existing annotation results according to the known miRNA >piRNA> rRNA > tRNA > snRNA > sno RNA > novel miRNA priority to ensure that each small RNA has a unique annotation.
4.Expression level analysis  The Reads Count value of the miRNA is counted based on the number of sequences aligned to the mature miRNA. The first abundance in the miRNA of the same name was chosen as the abundance of the miRNA for subsequent analysis.
5.Differential expression analysis  DESeq v 1.30.0) was condu cted to analyze the differential expressed miRNAs, tr anscripts with |log2FoldChange|>1 and P value<0.05 were considered as differentially expressed miRNAs.
Assembly: hg38
Supplementary files format and content: Matrix table with Counts per million  values for each Sample

source_name: Plasma

extract_protocol:
Total RNA was isolated using the Trizol Reagent (Invitrogen Life Technologies), after which the concentration, quality and integrity were determined using a NanoDrop spectrophotometer (Thermo Scientific）
Small RNA libraries were constructed using the NEBNext Multiplex Small RNA Library Prep Set for Illumina (New England Biolabs, Inc.) according to the manufacturer’s instructions.

value: Plasma


title: WT_F_Apc_2


title: patient9

description: exosomal microRNAs


title: WT_M_Apc_1


title: HEK293CDK7as_MNaseChIPseq_PolII_RPB1_DMSO_30min_Rep2


title: MCF10A_Unc_2


title: MCF10A_Unc_1


title: MCF10A_ECM_1


title: patient8

description: exosomal microRNAs


title: MCF10A_ECM_2


title: C_M_Apc_3

value: control


title: HEK293CDK7as_MNaseChIPseq_PolII_RPB1_DMSO_30min_Rep1


title: HEK293CDK7as_MNaseseq_1-NM-PP1_15min_Rep1

value: 15 min 1-NM-PP1

value: MNase-seq


title: patient7

description: exosomal microRNAs


title: BR Inoculated

city: Mysuru

first: Chandrakanth

last: R

state: Karantaka

address: JLB Road

department: Molecular Biology

country: India

institute: Yuvaraja's College, University of Mysore

data_processing:
2x100 paired-end sequence reads
Raw reads in the form of fastq files were analysed using FastQC tool, for quality check  based on per sequence (04 parameters) and per base (05 parameters analysis vide different parameters like read length, phred score, GC content and ambiguous bases using CLC bio Genomic Workbench version 9.
Reads were trimmed to remove the low quality bases at 5' and 3' ends. Only reads greater than Q30 have been selected for the further downstream analysis.
High quality clean reads were mapped to the rice reference genome RGAP 7 using reference assembly tool of CGWB
The mapped read data was summarised into a matrix for each sample over count parameters. Subsequently count data was compared between the samples to identify the differentially expressed genes and were further filtered based on fold change, p value and FDR p value.
Supplementary_files_format_and_content: tab-delimited text files include count data for each Sample.

organism: Oryza sativa

source_name: Leaves

growth_protocol: Plants were grown on greenhouse conditions like 14 h light and 10 h dark with optimum temperature of about 24± 2°C and high realtive humidity (90%).

extract_protocol:
Rice leaves were collected, flash frozen on ice and preserverd in RNAlater solution and RNA was harvested using RNeasy Plant Mini Kit (Qiagen). 1µg of Total RNA was taken for mRNA library preparation using NEBNext® Ultra™ RNA Library Prep Kit.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: M.oryzae Conidia concentration of about 105 conidia/ml mixed with 0.1% Tween-20 suspension solutiuon was sprayed during evening hour and plants were kept in complete dark for first 24 hrs.

tag: cultivar

value: Leaves


title: HEK293CDK7as_MNaseseq_1-NM-PP1_15min_Rep2

data_processing:
All sequencing data were demultiplexed with Illumina bcl2fastq demultiplexing software, provided the barcodes for each of the samples, and subsequently quality checked with FastQC (Babraham-Institute) (Andrews, 2010).
TT-seq: For all samples, paired-end 75 base reads were mapped using STAR (v 2.6.1b) (Dobin et al., 2013) to the GRCh38.p13 genome assembly merged with synthetic RNA spike-in sequences (Schwalb et al., 2016; Żylicz et al., 2019) with a maximum 2% mismatches [mapping parameters: --outFilterMismatchNoverLmax 0.02 --outFilterMultimapScoreRange 0 --alignIntronMax 500000]. SAMtools (v 1.3.1) (Li et al., 2009) was used to quality filter SAM files, whereby alignments with MAPQ smaller than 7 (-q 7) were skipped, and only proper pairs (-f2) were selected.  A spike-in (RNA) normalization strategy was used essentially as described (Schwalb et al., 2016). This spike-in normalization allows for calculating sequencing depth σ_j, cross-contamination rate ϵ_j, and antisense bias ratio c_j. Sequencing depth σ_j allows to observe global shifts. The read counts (kij) for spike-ins were calculated using HTSeq (Anders et al., 2015).
mNET-seq: For all samples, paired-end 45 base reads were trimmed for adapters (-a TGGAATTCTCGGGTGCCAAGG -A GATCGTCGGACT), and low-quality bases (< Q20) were removed with Cutadapt (v 1.9.1) (Martin, 2011). The reads were then mapped to the GRCh38.p13 genome assembly merged with the yeast genome version SacCer3 using STAR (v 2.5.2b) (Dobin et al., 2013) with a maximum of 2% mismatches [mapping parameters: --outFilterMultimapScoreRange 0 --outFilterMismatchNoverLmax 0.02 --outFilterMatchNmin 16 --outFilterScoreMinOverLread 0  --outFilterMatchNminOverLread 0 --alignIntronMax 500000 ]. Samtools (v 1.3.1) (Li et al., 2009) was used to quality filter SAM files, where alignments with MAPQ smaller than 7 (-q 7) were skipped, and only proper pairs (-f 2) were selected.   Read counts (kij) for yeast genes for each condition j (control or CDK7as inhibited) were calculated using HTSeq (Anders et al., 2015). The counts data was used to generate normalization factors with DESeq2 (Love et al., 2014) to normalize the mNET-seq data.
MNase-seq: For all samples, paired-end 45 base reads were trimmed for low-quality bases, random matches, and for a minimum length [trimming parameters: -q 20,20 -O 12 -m 25] with Cutadapt (v 1.9.1) (Martin, 2011). Processed reads were then mapped to the GRCh38.p13 genome assembly using STAR (v 2.6.1b) (Dobin et al., 2013) with maximum 2 percent mismatches [mapping parameters: --outFilterMismatchNoverLmax 0.02 --outFilterMultimapScoreRange 0 --alignIntronMax 1]. Samtools (v 1.3.1) (Li et al., 2009) was used to quality filter SAM files, where alignments with MAPQ smaller than 7 (-q 7) were skipped, and only proper pairs (-f 2) were selected.
MNase-ChIP-seq: For all samples, paired-end 43 base reads were mapped using Bowtie 2 (v 2.3.4.1) (Langmead and Salzberg, 2012) to the merged human hg38 (GRCh38.p13) and Drosophila (BDGP6.28) genome assembly [mapping parameters: --no-discordant --no-mixed --very-sensitive]. SAMtools (v 1.3.1) (Li et al., 2009) was used to quality filter SAM files, whereby alignments with MAPQ smaller than 7 (-q 7) were skipped, and only proper pairs (-f2) were selected. Further data processing was carried out using the R/Bioconductor environment. Duplicate fragments and fragments longer than 100 bp (70 bp for Pol II) were excluded from further analysis. MNase-ChIP-seq coverages were obtained from piled-up fragment midpoint counts for every genomic position. Pol II, Ser5P, and SPT5 coverages were normalized to the signal at untranscribed regions to allow for the detection of global changes. To this end, genome-wide coverage was calculated from all MNase-ChIP-seq fragment midpoints in consecutive 2 kbp bins throughout the non-transcribed human genome (i.e. excluding 32,789 transcribed units we annotated using GenoSTAN (Zacher et al., 2017) and everything annotated in GENCODE v31) and used to generate normalization factors with DESeq2 (Love et al., 2014). For all other factors, which exhibited a peak-like binding behavior, normalization factors were obtained from total human fragment counts. Bigwig files are for normalized (except Inputs), entire fragment coverages.
Assembly: GRCh38
Supplementary files format and content: bigwig files

extract_protocol:
TT-seq: Total RNA was extracted using TRIzol reagent (Invitrogen, 15596026), 4sU-labeled RNA was extracted according to Gressel at al., protocols.io, 2019.
mNET-seq: mNET-seq was performed as previously described (Caizzi et al. 2021; Gressel et al. 2017; Nojima et al. 2016; Schlackow et al. 2017) with minor changes. 50 mio CDK7as cells were treated for 15 or 30 min with 7.5 μM of 1-NM-PP1 or DMSO (solvent control). After the treatment, chromatin isolation was performed as described in the protocol for cellular fractionation (Conrad et al. 2017) using 25 mio cells per reaction. All buffers were supplemented with 7.5 μM 1-NM-PP1 or respective amounts of DMSO, protease (Sigma-Aldrich, P8340), and phosphatase (Roche, 4906837001) inhibitors. After the fractionation, the chromatin pellet was digested in 100 μl MNase digest buffer (1x MNase buffer, 1x BSA, 100 mM NaCl, 50 units/μl MNase (NEB, M0247S)) at 37°C and 1,400 rpm for 2 min. The digestion was stopped by adding 11 μl of 250 mM EGTA, and samples were centrifuged at 13,000 g for 5 min at 4°C. Supernatants were pooled together for each sample and 8-fold diluted with IP buffer (50 mM Tris HCl pH 7.4, 150 mM NaCl, 0.05% NP-40, 0.3% empigen (Sigma-Aldrich, 30326)). 15 μg of anti-Pol II CTD antibody (Diagenode, C15200004) conjugated with 125 μl of Dynabeads M-280 Sheep Anti-Mouse IgG (Invitrogen, 11201D) were added to each sample and incubated on a rotating wheel at 4°C and 12 rpm for 1 h. After IP, beads were washed seven times with 1 ml of IP buffer and once with 300 μl of PNKT buffer (1x T4 PNK buffer (NEB, M0236L), 0.1% Tween-20). After washes, 5’ RNA ends were phosphorylated with T4 Polynucleotide Kinase (PNK) (3' phosphatase minus; NEB, M0236L) at 800 rpm for 10 min at 37°C. Next, beads were washed once with IP buffer, and 0.5 ml of TRIzol reagent (Invitrogen, 15596026) was added to the beads. 2.5 ng of S. cerevisiae spike-ins were added to each sample in TRIzol. RNA spike-ins for mNET-seq were obtained from S. cerevisiae cells as previously described (Caizzi et al. 2021). RNA was extracted from beads, followed by RNA precipitation with 100% ethanol and GlycoBlue coprecipitant (Invitrogen, AM9515) at -80°C overnight. RNA pellets were washed twice with 70% ethanol, air-dried, and dissolved in 7 M urea. In order to size select 25-110 nt RNA, samples were resolved on 6% polyacrylamide gel containing 7 M urea. RNA of corresponding sizes was extracted from the gel and precipitated with 100% ethanol and GlycoBlue coprecipitant.
MNase-seq: 5 mio CDK7as cells were treated for 15 min with 7.5 μM of 1-NM-PP1 or DMSO (solvent control) and centrifuged at 500 g for 3 min at 4°C. Cell pellets were resuspended in 50 μl of Buffer I (10 mM Tris-HCl pH 7.5, 10 mM NaCl, 2.5 mM MgCl2, 0.5% NP-40, 10 % sucrose, 0.5 mM CaCl2) and incubated at 37°C for 2 min. Next, we added 500 μl of Buffer II (50 mM Tris-HCl pH 7.5, 50 mM NaCl, 2.5 mM MgCl2, 0.5% NP-40, 10 % sucrose, 2 mM CaCl2) supplemented with 3,000 U of MNase (NEB, M0247S) and incubated at 37°C for 9 min. MNase digestion was quenched with 11 μl of EGTA (500 mM). 550 μl of SDS Lysis Buffer (50 mM Tris-HCl pH 8.0, 1% SDS, 10 mM EDTA) was added, and samples were incubated on ice for 10 min. Samples were then treated with 0.1 μg/μl RNAse A (Thermo Scientific, EN0531) for 1.5 h at 37°C and 0.05 μg/μl Proteinase K (Life Technologies, AM2546) for 4 h at 65°C. DNA was extracted with an equal volume of phenol/chloroform/isoamyl alcohol (Sigma-Aldrich, P2069), followed by DNA precipitation with an equal volume of 100% ethanol and 200 mM NaCl for 2 h at -20°C. DNA pellets were washed twice with 1 ml of 70% ethanol, air-dried, and dissolved in water.
MNase-ChIP-seq: MNase-ChIP-seq was performed as previously described (Skene et al. 2015) with the following changes. 40 mio CDK7as cells were treated for 30 min with 7.5 μM of 1-NM-PP1 or DMSO (solvent control), following 8 min crosslinking with 1% formaldehyde (16% formaldehyde (w/v), methanol-free, Thermo Fisher Scientific, 28908). The crosslinking was quenched for 5 min with 125 mM glycine, and cells were washed with 10 ml PBS at room temperature. 2 ml of ice-cold PBS was added to plates for cell scraping. Scraped cells were transferred to falcons, washed twice with 5 ml ice-cold PBS, and centrifuged at 200 g for 5 min at 4°C. All buffers were supplemented with protease (Sigma-Aldrich, P8340) and phosphatase (Roche, 4906837001) inhibitors. Cell pellets were lysed with 5 ml of Farnham Lysis buffer (5 mM Pipes pH 8, 85 mM KCl, 0.5% NP-40) for 10 min on ice. Nuclei were pelleted by centrifugation at 1,700 g for 5 min at 4°C and washed with ice-cold PBS. Nuclei pellets were resuspended in 200 μl of 1% SDS Lysis buffer (50 mM Tris HCl pH 8.0, 10 mM EDTA, 1% SDS) and incubated for 10 min at room temperature to permeabilize nuclei. The reaction was quenched with 1.8 ml of IP buffer (20 mM Tris HCl pH 8.0, 150 mM NaCl, 1 mM EDTA, 1% Triton X-100) supplemented with 5 mM CaCl2 and incubated for 2 min at 37°C before the addition of 16,000U of MNase (NEB, M0247S) for 20 min at 37°C. MNase digestion was quenched with 40 μl of EDTA (500 mM) and 20 μl EGTA (500 mM). 1 ml of sample was transferred to AFA milliTubes (Covaris, 520135) and sonicated with a Covaris S220 Focused-ultrasonicator with the following parameters: 5% duty cycle, 140 W peak incident power, 200 cycles per burst, 600 sec processing time, 6°C bath temperature, continuous degassing mode, water level 8. Sonicated samples were centrifuged at 10,000 g for 15 min at 4°C, and supernatants were snap-frozen in liquid nitrogen and stored at −80°C before use. For each IP, 30 μg of sample chromatin and 200 ng of Drosophila S2 MNase-digested crosslinked chromatin were diluted in IP buffer to obtain a final concentration of 0.05% SDS and a final volume of 1.6 ml. Pre-clearing was carried out by adding 20 μl of Dynabeads Protein G (Thermo Fisher Scientific, 10009D) diluted in IP buffer, and incubating for 30 min at 4°C on the rotation wheel. Pre-cleared samples were incubated with 10 μg of Pol II (RPB1, Diagenode, C15200004), 10 μg of Ser5P (phospho-CTD Ser-5 of RPB1, Merck Millipore, 04-1572), 8 μl of Mediator (MED26, Cell Signaling, 14950), 6 μg of TFIIB (Santa Cruz, sc-271736), 5 μg of TFIIE (TFIIE-α, abcam, ab28177), 6 μg of NELF (NELF-E, Bethyl Laboratories, A301-913A) or 6 μg of SPT5 (Proteintech, 16511-1-AP) together with 1 μg of Spike-in (Active Motif, 61686) antibodies at 4°C overnight. The next day samples were centrifuged at 16,000g for 2 min at 4°C and inputs were taken for each sample. Samples were incubated for 2 h at 4°C with Dynabeads Protein G. Beads were washed once with Buffer 1 (20 mM Tris HCl pH 8.0, 150 mM NaCl, 2 mM EDTA, 1% Triton X-100, 0.1% SDS), four times with Buffer 2 (20 mM Tris HCl pH 8.0, 500 mM NaCl, 2 mM EDTA, 1% Triton X-100, 0.1% SDS), once with Buffer 3 (10 mM Tris HCl pH 8.0, 250 mM LiCl, 1 mM EDTA, 1% NP-40, 1% sodium-deoxycholate), and three times with TE buffer (10 mM Tris HCl pH 8.0, 50 mM NaCl, 1 mM EDTA). After the last wash, 200 μl of elution buffer (0.1 M NaHCO3, 160 mM NaCl, 1% SDS) was added to the beads, and samples were de-crosslinked overnight at 65°C. Next, samples were diluted with 200 μl of PBS and treated with 0.02 μg/μl RNAse A (Thermo Scientific, EN0531) for 1.5 h at 37°C and 0.2 μg/μl Proteinase K (Life Technologies, AM2546) for 2 h at 45°C. DNA was extracted with an equal volume (400 μl) of phenol/chloroform/isoamyl alcohol (Sigma-Aldrich, P2069), followed by DNA precipitation with 100% ethanol and 200 mM NaCl for 1 h at -80°C. DNA pellets were washed twice with 70% ethanol, air-dried, and dissolved in water.
TT-seq: Libraries were prepared from 100 ng of RNA with the strand-specific Ovation Universal RNA-Seq library kit (NuGEN, 0343-32).
mNET-seq: 8.4 ng of RNA for replicate 1 and 12.3 ng of RNA for replicate 2 were used for library preparation with TruSeq Small RNA Library Prep Kit (RS-200-0048) according to the manufacturer’s instructions.
MNase-seq: Libraries were prepared with the NEBNext Ultra II DNA Library Prep Kit for Illumina (NEB, E7645S) according to the manufacturer’s instructions.
MNase-ChiP-seq: Libraries were prepared with the NEBNext Ultra II DNA Library Prep Kit for Illumina (NEB, E7645S) according to the manufacturer’s instructions with adjusted for short fragments end repair, 5’ phosphorylation, and dA-tailing thermocycler program (30 min at 20°C, 1 h at 50°C; the heated lit set to 60°C). After amplification, libraries were double-sided (1.0 – 1.6x) size-selected with Agencourt AMPure XP beads (Beckman Coulter, A63881).

tag: cell line


title: BR Control


title: patient6

description: exosomal microRNAs


title: HEK293CDK7as_MNaseseq_DMSO_15min_Rep2

tag: cell type

value: 15 min DMSO


title: C_M_Apc_1


title: C_F_Apc_3


title: HR Control


title: patient4

description: exosomal microRNAs


title: C_M_Apc_2


title: HEK293CDK7as_MNaseseq_DMSO_15min_Rep1

value: genome edited HEK293

tag: genotype


title: C_F_Apc_2

tag: treatment


title: S2_Pol_II_2

first: Xiangyi

last: Shen

address: D303 Zhengyutong Medical Building, Tsinghua University

department: The Institute for Immunology, School of Medicine

institute: Tsinghua University

data_processing:
Raw reads in FASTQ format were processed through fastp (v0.19.11) to remove adapter-containing, poly-N containing, or low-quality reads in order to generate clean data.
Index of the reference genome (mm10) was built using BWA (v 0.7.12), and clean reads were aligned to the reference genome using BWA-MEM(v 0.7.12).
The bamCoverage tool (v2.2) of Deeptools was used to generate bigWig files using CPM normalization.
Assembly: mm10
Supplementary files format and content: bigWig
Library strategy: CUT&Tag

source_name: Common dendritic cell progenitor

extract_protocol:
CD11c–CD127–CD117intFLT3+ CDPs were sorted by flow cytometry from the bone marrow of wild-type C57BL/6 mice.
1×10^5 CDPs/sample were bound to beads and were subjected for immunoprecipitation with primary TRIM33, CDK9, S2 Pol II antibody or control IgG at 4 °C overnight. A secondary anti-rabbit antibody was then added and incubated under gentle agitation at room temperature  for 1 hr. Cells were then washed and incubated in Dig-300 buffer with 0.04 μM hyperactive pG-Tn5 transposon at RT for 1 hr. Cells were washed and then incubated in 100 μl of tagmentation buffer (10 mM MgCl2 in Dig-300 buffer) at 37 °C for 1 hr. Reaction was stopped by addition of SDS, EDTA and proteinase K and incubated at 55 °C for 1 hr and then DNA were extracted. For library amplification, 24 μL of DNA was mixed with 10 μL 5×TruePrep Amplify Buffer (TAB), 1 μL TAE, and 5 μL barcoded i5 and i7 primers and amplified for 20 cycles. PCR products were purified with VANTS DNA Clean beads (Vazyme). Libraries were sequenced on an Illumina HiSeq X Ten platform.

tag: antibody

value: Common dendritic cell progenitor

value: wild-type

value: untreated


title: HEK293CDK7as_mNETseq_1-NM-PP1_30min_Rep2

first: Michael

last: Lidschreiber

institute: Max-Planck-Institute for Multidisciplinary Scienes

value: Human embryonic kidney

value: analog-sensitive CDK7

tag: immunoprecipitation antibody

tag: method

value: mNET-seq


title: patient3

description: exosomal microRNAs

tag: tissue


title: HR Inoculated


title: C_F_Apc_1


title: S2_Pol_II_1

tag: strain


title: B_M_Apc_3

value: black rice


title: patient2

description: exosomal microRNAs


title: patient5

description: exosomal microRNAs


title: HEK293CDK7as_mNETseq_1-NM-PP1_15min_Rep2


title: patient1

description: exosomal microRNAs


title: B_M_Apc_2


title: TRIM33_2


title: HEK293CDK7as_mNETseq_1-NM-PP1_15min_Rep1


title: HEK293CDK7as_mNETseq_1-NM-PP1_30min_Rep1

value: 30 min 1-NM-PP1


title: B_F_Apc_3


title: TRIM33_1


title: K28PE-WT-IP

city: Nanjing

first: Yuanjiao

state: China

address: Xianlin Road

institute: nanjing university of chinese medicine

data_processing:
Fastp 0.19.11 software performs basic quality statistics for raw reads
QC analysis was used by  fastqc v0.11.5
burrows Wheeler aligner(0.7.12-r1039) can  compare reads to reference genome
Peak calling was performed by MACS22.1.0
The motifs were recognized by Homer findMotifsGenome.pl(v4.9.1)
Assembly: GRCh38

source_name: Multiple Myeloma cells

growth_protocol: Human multiple myeloma cell lines K28MS-PE were cultured in RPMI 1640 Medium  with 10% fetal bovine serum 100 U/ml penicillin and 100 mg/ml streptomycin. All cells were cultured in a humidified atmosphere of 5 % CO2 at 37℃.

extract_protocol:
anti-ac4C antibody (1 μg) or rabbit monoclonal IgG Isotype control (1 μg) were pre-coupled to 300 μg Protein G Dynabeads (ThermoFisher Scientific) in PBS for 1 hr at room temperature. DNase-treated total RNA (1 μg) from HeLa cells was spiked with 10 pg (1:10−5), 1 pg (1:10−6) or 0.1 pg (1:10−7) of ac4C- or CRNA and immunoprecipitated for 4 hr at 4 °C in 100 μl of acRIP buffer containing PBS, 0.05% Triton X-100, 0.1 % BSA, 40U murine RNase inhibitor (NEB) and anti-ac4C or IgG pre-coupled Protein G Dynabeads. After immunoprecipitations, beads were washed five times in acRIP buffer and elution of RNA was carried out by RNase-free Proteinase K (50 μg, ThermoFisher Scientific) digestion in 100 μl buffer containing 50 mM Tris-HCl [pH7.5], 75 mM NaCl, 6.25 mM EDTA and 1 % SDS for 1h at 37 °C. RNA was extracted by Phenol:Chloroform [pH 4.5] and ethanol precipitation using 0.3M sodium acetate [pH 5.5] and 15 μg/mL linear acrylamide.
Acetylated RNA immunoprecipitation and sequencing (acRIP-seq) Poly(A) RNA from parental and NAT10−/− was used for acRIP-seq analysis. To evaluate clones representing distinct residual ac4C levels, in replicate 1, the NAT10−/− RNA sample corresponded to the pool of NAT10−/−A and NAT10−/−C. In replicate 2, the NAT10−/− sample corresponded only to NAT10−/−A. Poly(A) RNA was isolated by two rounds of oligod(T) selection and fragmented using NEBNext® Magnesium RNA Fragmentation buffer for 5 min at 94 °C. Eight picograms of ac4C-RNA probe was spiked into 8 μg of fragmented poly(A) RNA followed by immunoprecipitation with 1 μg anti-ac4C or IgG pre-coupled to Protein G Dynabeads as described above (see Acetylated RNA immunoprecipitation). Illumina libraries were constructed for inputs (parental vs. NAT10−/−, replicate 1 and 2), acRIPs (parental vs. NAT10−/−, replicate 1 and 2), and IgG (parental, replicate 1) using the NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina®. Libraries were multiplexed using TruSeq V4.0 chemistry and sequenced for 126 cycles in paired-end mode

tag: genotype/variation

value: WT

value: anti-ac4C antibody


title: HEK293CDK7as_mNETseq_DMSO_30min_Rep2

city: Goettingen

address: Am Fassberg 11

value: 30 min DMSO

value: Pol II CTD antibody (Diagenode, C15200004)


title: B_M_Apc_1

value: intestinal tract


title: K28PE-OE-IP

value: OE


title: IgG

value: IgG


title: IP-Naive-rep3

first: Shuyang

last: Yu

address: 2 Yuanmingyuan West Road

department: College of Biological Sciences

institute: China Agricultural University

data_processing:
FastQC was used to identify features of the raw data that may indicate quality problems
Trimmomatic was used to trim raw fastq files
Kallisto was used to mapped to GRCm38 cDNA sequences
DESeq2 was used to calculate differential expression of m6A levels for each gene
Assembly: mm10
Supplementary files format and content: xls format. Read counts for each gene

source_name: Spleen

extract_protocol:
Wild-type naïve (CD44-CD62L+) CD8+ T cells, effector  (CD44+CD62L-) CD8+ T cells and memory  (CD44+CD62L+) CD8+ T cells were sorted. Total RNAs were extracted from indicated cells and then submitted to SLIM-Seq protocol.
SLIM-Seq data were analyzed as previously described (Rong Yin et al., 2022)

value: Spleen

value: naive CD8 (CD44-CD62L+CD8+)

value: 6~8 weeks


title: HEK293CDK7as_mNETseq_DMSO_30min_Rep1

value: genome edited HEK293


title: B_F_Apc_2


title: B_F_Apc_1


title: CDK9

value: Common dendritic cell progenitor

value: wild-type

value: untreated


title: K28PE-OE-input

value: input


title: HEK293CDK7as_mNETseq_DMSO_15min_Rep2

value: Human embryonic kidney

value: analog-sensitive CDK7

value: 15 min DMSO

value: mNET-seq


title: K28PE-WT-input

value: WT


title: IP-Naive-rep2


title: HEK293CDK7as_mNETseq_DMSO_15min_Rep1

value: Pol II CTD antibody (Diagenode, C15200004)


title: HEK293,RNaseR,nanopore,rep3

city: Ipswich

first: Zhiyi

last: Sun

state: MASSACHUSETTS

address: 240 County Road

department: Research

institute: New England Biolabs

data_processing:
PacBio consensus reads were generated using the SMRT tools (version 10.2.0.133434). Raw nanopore data were basecalled in high accuracy mode with Dorado 7.0.9 and filtered by average quality (≥ 7).
Sequencing reads in FASTQ format from either PacBio or Nanopore were first mapped to the rDNA reference sequences using minimap2 and the unmapped reads were kept for downstream circRNA analysis.
Then circular consensus sequences and candidate circRNAs were identified using the CIRI-long bioinformatics pipeline. We applied an additional filter to remove consensus sequences that were called from any read that contains fewer than 2 copies of repeat sequences. Candidate circRNA sequences were identified based on the GRCh38 reference genome sequence and annotation. Then all candidate circRNA sequences from the subsets were aggregated together using the CIRI-long collapse program to determine BSJ sites and calculate the relative isoform usage index of each BSJ.
Finally, for each BSJ, the read counts of its isoforms were calculated from the isoform usage index (*collapse.isoforms file) and the total read counts of that BSJ as isoform expression = isoform usage index * BSJ read counts.
Assembly: GRCh38/hg38
Supplementary files format and content: Tabular data file with circRNA isoform counts
Library strategy: Induro-RT mediated circRNA-sequencing (IMCR-seq)

extract_protocol:
NEB Monarch Total RNA Miniprep Kit was used for home made HEK293 cell RNA, guanidine thiocyanate tecnniques were reported to be used for commercially obtained RNA and Quick-cfRNA Serum & Plasma Kit from Zymo Research was used for RNA extraction from plasma
Total RNA or rRNA depleted, RNaseR treated RNA were reverse transcribed with a group II intron RT then the cDNAs were sized selected with SPRI beads. Second strand synthesis and amplification were done with Phi29 DNA polymerase. T7 endonuclease I was used to digest the branched DNA into double stranded DNA and ONT and PacBio library preparation kits for dsDNA were used to make libraries for sequencing.


title: hMSCs SiO2 1

city: Bryan

first: Irtisha

last: Singh

state: TX

address: 8447 Riverside Pkwy Medical Research and Education Building II

institute: Texas A&M Health Science Center

description: SiO2 treatment

data_processing:
Sequence cluster identification, quality prefiltering, base calling and uncertainty assessment were done in real time using Illumina's NCS 1.0.2 and RFV 1.0.2 software with default parameter settings. Sequencer .cbcl basecall files were demultiplexed and formatted into .fastq files using bcl2fastq 2 2.19.0 script configureBclToFastq.pl.
Sequenced reads were trimmed for adaptor sequence and aligned by STAR aligner
Fragments Per Kilobase of exon per Megabase of library size (FPKM) were calculated. In short, exons from all isoforms of a gene were merged to create one meta-transcript. The number of reads falling in the exons (excluding 5UTR and 3UTR) of this meta-transcript were counted and normalized by the size of the meta-transcript and by the size of the library (only reads mapping to annotated regions of the genome were used for the total libraray size).
Genome_build: hg38
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for all samples

source_name: Mesenchymal stem cells

growth_protocol: hMSCs were cultured under media conditions consisting of α-minimal essential media (alpha-MEM, Hyclone, GE Sciences) with 16.5% fetal bovine serum (Atlanta Biologicals, USA) supplemented with 10 mM β-glycerophosphate (Sigma-Aldrich) and 50 µM ascorbic acid (BDH Chemicals)) and 1% penicillin/streptomycin (100 U/100 µg/mL, Gibco). After every 2-3 days, half of culture media was exchanged for fresh media. Cells were passaged with 0.5% trypsin-EDTA upon reaching confluency of ~70% and seeded at ~2500 cells/cm2. All experiments were completed with cell populations under P5.

extract_protocol:
At the end of 21 days of culture, total RNA was extracted (Zymogen, High Purity RNA Isolation kit), and then converted into cDNA (>1 μg).
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: hMSCs were cultured and treated with/without nanomaterials (100µg/mL) nSi, nHA, nWH, or nSiO2

value: Bone marrow derived mesenchymal stem cells


title: IP-Naive-rep1


title: HEK293CDK7as_TTseq_TotalRNA_1-NM-PP1_60min_Rep2

value: 60 min 1-NM-PP1, 10 min 4sU

value: TT-seq


title: HEK293,RNaseR,nanopore,rep2


title: IP-memory-rep3

value: Spleen

value: Memory CD8 (CD44+CD62L+CD8+)


title: hMSCs SiO2 2

description: SiO2 treatment


title: HEK293CDK7as_TTseq_TotalRNA_1-NM-PP1_60min_Rep1


title: hMSCs WH 2

description: WH treatment


title: HEK293,RNaseR,nanopore,rep1


title: IP-memory-rep2

value: 6~8 weeks


title: HEK293CDK7as_TTseq_TotalRNA_1-NM-PP1_30min_Rep2

value: genome edited HEK293

value: 30 min 1-NM-PP1, 10 min 4sU


title: hMSCs WH 1

description: WH treatment


title: HEK293,total,nanopore,rep2


title: IP-memory-rep1


title: hMSCs nHA 2

description: nHA treatment


title: HEK293CDK7as_TTseq_TotalRNA_1-NM-PP1_30min_Rep1


title: HEK293,total,nanopore,rep3


title: IP-effector-rep3

value: effector CD8 (CD44+CD62L-CD8+)


title: hMSCs nHA 1

description: nHA treatment


title: HEK293CDK7as_TTseq_TotalRNA_1-NM-PP1_15min_Rep2

value: 15 min 1-NM-PP1, 10 min 4sU


title: HEK293,total,nanopore,rep1


title: HEK293CDK7as_TTseq_TotalRNA_1-NM-PP1_15min_Rep1


title: IP-effector-rep2


title: brain,RNaseR,nanopore,rep3

source_name: brain

value: brain


title: IP-effector-rep1


title: DLS490, SE, biol rep 3

city: Chapel Hill

first: Robert

last: Dowen

address: 321 Fordham Hall, CB7100

institute: University of North Carolina at Chapel Hill

description:
Column names listed in line one for each txt file
FC_gene_read_counts_Hh_study.txt
DESeq2_norm_gene_counts_Hh_study.txt
DESeq2_DEGs_DLS490_v_N2_Hh_study.txt
DESeq2_DEGs_DLS490_v_MT333_Hh_study.txt
DESeq2_DEGs_DLS487_v_DLS490_Hh_study.txt

data_processing:
The adapter-trimmed mRNA-Seq reads were mapped to the WS260 genome using the STAR aligner. The mapping was performed using a GTF obtained from Wormbase (c_elegans.PRJNA13758.WS260.canonical_geneset.gtf) with the parameter --outFilterIntronMotifs RemoveNoncanonical.
The gene-level read counts were calculated using the featureCounts algorithm of the subread package (Liao et al., Bioinformatics, 2014), using the following flags: -t exon -g gene_id -a c_elegans.PRJNA13758.WS260.canonical_geneset.gtf.
Differential expression was calculated relative to the N2 wild-type sample using the DESeq2 package using a 1% FDR cutoff (Love et al., 2014). Comparisons between individual mutants were also performed.
Assembly: WS260
Supplementary files format and content: FC_gene_read_counts_study_name.txt: A tab-delimited text file for each sub-study that includes gene level unnormalized read counts for each sample in matrix format generated using the featureCounts aligorithm.
Supplementary files format and content: DESeq2_norm_gene_counts_study_name.txt: A tab-delimited text file that for each sub-study includes gene level normalized read counts for each sample in matrix format generated using the DESeq2 aligorithm.
Supplementary files format and content: DESeq2_DEGs_sample1_v_sample2_study_name.txt: A tab-delimited text file that includes a list of differentially expressed genes, the log2 FC values, and the p values called by DESeq2 at a 1% FDR threshold for sample1 relative to sample2 for each sub-study.

source_name: Whole animals

extract_protocol:
Animals were harvested in M9 buffer, washed three times, and flash frozen in liquid Nitrogen. Total RNA was isolated using Trizol.
mRNA-Seq libraries were prepared using the Illumina TruSeq RNA Sample Preparation v2 kit. For the tra-1 and tir-1 studies, mRNA-Seq libraries were prepared by Novogene Inc.

value: rict-1(mg360) II

value: Standard growth conditions at 20C on NGM plates containing E. coli OP50

tag: developmental stage

value: Day 1 adults

value: Whole animals

tag: Sex

value: hermaphrodite


title: DLS490, SE, biol rep 2


title: HEK293CDK7as_TTseq_TotalRNA_DMSO_60min_Rep2

value: Human embryonic kidney

value: analog-sensitive CDK7

value: 60 min DMSO, 10 min 4sU


title: DLS490, SE, biol rep 1


title: HEK293CDK7as_TTseq_TotalRNA_DMSO_30min_Rep2

value: 30 min DMSO, 10 min 4sU

value: TT-seq


title: DLS487, SE, biol rep 3

description:
Column names listed in line one for each txt file
FC_gene_read_counts_Hh_study.txt
DESeq2_norm_gene_counts_Hh_study.txt
DESeq2_DEGs_DLS487_v_N2_Hh_study.txt
DESeq2_DEGs_DLS487_v_MT333_Hh_study.txt
DESeq2_DEGs_DLS487_v_DLS490_Hh_study.txt


title: input-Naive-rep1

value: Spleen

value: naive CD8 (CD44-CD62L+CD8+)

value: WT

tag: age


title: brain,RNaseR,nanopore,rep1


title: HEK293CDK7as_TTseq_TotalRNA_DMSO_60min_Rep1

value: genome edited HEK293


title: input-memory-rep3

value: Memory CD8 (CD44+CD62L+CD8+)

value: 6~8 weeks


title: HEK293CDK7as_TTseq_TotalRNA_DMSO_30min_Rep1


title: HEK293CDK7as_TTseq_TotalRNA_DMSO_15min_Rep2

value: 15 min DMSO, 10 min 4sU


title: DLS487, SE, biol rep 1


title: input-Naive-rep3


title: DLS440, SE, biol rep 3

description:
Column names listed in line one for each txt file
FC_gene_read_counts_lin-29_study.txt
DESeq2_norm_gene_counts_lin-29_study.txt
DESeq2_DEGs_DLS440_v_N2_lin-29_study.txt
DESeq2_DEGs_DLS440_v_MT333_lin-29_study.txt
DESeq2_DEGs_DLS439_v_DLS440_lin-29_study.txt


title: brain,total,nanopore,rep1

value: brain


title: input-memory-rep1


title: HEK293CDK7as_TTseq_LabeledRNA_1-NM-PP1_60min_Rep2

value: 60 min 1-NM-PP1, 10 min 4sU


title: DLS440, SE, biol rep 2

value: Standard growth conditions at 20C on NGM plates containing E. coli OP50

value: Day 1 adults


title: brain,RNaseR,nanopore,rep2


title: DLS440, SE, biol rep 1

value: Whole animals

value: hermaphrodite


title: brain,total,nanopore,rep2


title: DLS439, SE, biol rep 3

description:
Column names listed in line one for each txt file
FC_gene_read_counts_lin-29_study.txt
DESeq2_norm_gene_counts_lin-29_study.txt
DESeq2_DEGs_DLS439_v_N2_lin-29_study.txt
DESeq2_DEGs_DLS439_v_MT333_lin-29_study.txt
DESeq2_DEGs_DLS439_v_DLS440_lin-29_study.txt


title: input-effector-rep1

value: effector CD8 (CD44+CD62L-CD8+)