title: Mrps5 fl/fl; MCM mouse heart tissue, cKO, 12 weeks after tamoxifen injection, rep 3

value: Heart

value: Mrps5 fl/fl; MCM

value: Tamoxifen injection


title: iN_46_cre_d7_rep2

status: Public on Feb 14 2023

type: SRA

city: Heidelberg

first: Bryce

last: Lim

address: In Neuenheimer Feld 280

country: Germany

institute: DKFZ

library_source: transcriptomic

data_processing:
Sequenced reads were trimmed for adaptor sequence and masked for low-complexity or low-quality sequence
Filtered reads were mapped to hg38 with STAR to generate raw counts files
Differential expression and normalised counts were determined using DESeq2 (R package version 1.28.1)
Genome_build: hg38
Supplementary_files_format_and_content: Tab-delimited text file with raw counts for all samples
Supplementary_files_format_and_content: Tab-delimited text file with DESeq2 output for all conditions (sample-collapsed)
Supplementary_files_format_and_content: Tab-delimited text file with normalised counts for all samples

molecule: total RNA

organism: Homo sapiens

source_name: NGN2-induced neurons

growth_protocol: H1 (male) human ES cells (WiCell) were maintained feeder-free in mTeSR Plus medium (Stem Cell Technologies) and one allele of human MYT1L (ENSG00000186487) was targeted by  homologous recombination to flank exon 7 with loxP sites. Cre-mediated recombination deleted exon 7 resulting in a frameshift starting in exon 8 at amino acid (aa) 19 and terminating in a premature STOP at aa 69 in exon 9 (p.V19AfsX51). Heterozygous targeting was confirmed by Sanger sequencing that showed insertion of loxP sites within one allele for each clone (+/fl) and MYT1L protein depletion was confirmed by Western blot directly in induced human neurons.

extract_protocol:
For human neurons pure cultures of induced human neurons at day 7 or induced human neurons co-cultured with primary mouse glia at day 43 were harvested by addition of TRIzol to the cultures (Invitrogen) and RNA was isolated using the RNA Miniprep kit (Zymo Research).
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: Lentivirus encoding doxycycline-inducible Ngn2 and either cre-recombinase or an inactive truncated delta-cre-recombinase, both under control of the ubiquitin promoter, were used to transduce conditional MYT1L knockout cells (+/fl). On day 0, Doxycycline (2 mg/ml, Sigma) was added to induce Ngn2 expression and retained in the medium until day 14. On day 1, cells were treated with Accutase (Gibco) and plated on Matrigel (Corning)-coated 6-well plates in DMEM/F12 containing N2-supplement, non-essential amino acids (all from Invitrogen), human BDNF (10 ng/ml, PeproTech), human NT-3 (10 ng/ml, PeproTech), and mouse Laminin-1 (0.2 mg/ml, Invitrogen) containing 10 µM Y-27632 (Axon Medchem) and puromycin (2 µg/ml c.f.) was added for 3 days to select infected cells. On day 4, iN cells were treated with Accutase and replated in Neurobasal-A medium supplemented with B27/glutamax (Invitrogen) containing BDNF, NT3, and Laminin-1, and Ara-C (2 µM, Sigma). For day 7 harvest, iN cells were seeded on Matrigel-coated 12-well plates without addition of glia. For functional maturation, iN cells were seeded on Matrigel-coated coverslips together with primary mouse glia with media changes every 2 days. After 1 week media was gradually exchanged with MEM supplemented with 0.5% glucose, 0.02% NaHCO3), 2 µM Ara-C (all Sigma), Transferrin (100 µg/ml c.f.; Merck), 5% FBS, B27, 0.5 mM L-glutamine (all Invitrogen), following media changes every 3–4 days for the duration of the experiment. All cells were grown at 37 °C and 5% CO2.

tag: donor cell line

tag: cell type

value: neuron

tag: harvest day

tag: treatment

value: cre

tag: clone

value: clone 2


title: mESC, STM2457, 6h, rep4

city: Würzburg

first: Julian

last: Konig

address: Am Hubland

institute: Würzburg University

data_processing:
Quality control: FastQC (v0.11.8)
Trimming: Cutadapt (v1.18) with non-default parameters: cutadapt -a file:NexteraPE-PE.fa --minimum-length 50 (additional parameter used for C643, HEK293T, RPE1, male mESC samples: --cut 1)
Mapping: STAR (v2.7.3a) with non-default parameters: STAR --runMode alignReads --outFilterMismatchNoverLmax 0.04 --outFilterMismatchNmax 999 --outStd SAM --outSAMattributes NH HI AS nM MD --outSJfilterReads Unique --outSAMunmapped Within --runThreadN 4 --readFilesCommand zcat --sjdbOverhang READLENGTH-1
Filter/remove secondary hits: Samtools (v1.9) with parameters: samtools view -F 256
Counts per gene: Subread (v2.0.0) with non-default parameters: featureCounts --donotsort -s2 -t exon (default used for -g gene_id)
Assembly: human: GRCh38.p12 (Gencode release 31) (+ ERCC spike-ins in C643, HEK293T, RPE1)
Assembly: mouse: GRCm38.p6 (Gencode release M23) (+ ERCC spike-ins in male mESC)
Supplementary files format and content: tab-delimited text (.tsv) files include absolute read counts per gene for each sample

molecule: polyA RNA

organism: Mus musculus

source_name: mESC

growth_protocol: For C643 and HEK293T: Cells were cultured in Dulbecco’s Modified Eagle Medium (DMEM, Life Technologies) supplemented with 10% fetal bovine serum (FBS, Life Technologies), 1% L-glutamine (Life Technologies) and 1% penicillin/streptomycin (P/S Life Technologies). For RPE1: Cells were cultured in DMEM/F12 (Life Technologies) supplemented with 10% foetal bovine serum (FBS, Life Technologies), 1% penicillin/streptomycin (P/S, Life Technologies), 1% L-Glutamin and 0.04% Hygromycin B (Life Technologies). Cells were grown at 37◦C with 5% CO2. For male and female mESC: Female mouse embryonic stem cells (mESC) were taken from a previous publication (PMID: 24506884). Single clones were picked and cultured until confluency. Male mESC were taken from a previous publication (PMID: 25569111).  Cells were cultured in 2i/LIF medium. Briefly, 500 ml medium was generated by adding 235 ml of each DMEM/F12 and neurobasal (Gibco; 21331020, 21103049), 7.5 ml BSA solution (7.5%, ThermoFisher; 11500496), 5 ml penicillin-streptomycin (Gibco; 15140-122), 2 mM L-Glutamin (ThermoSci; 25030024), 100 µM β-mercaptoethanol (Gibco; 21985023), 5 ml mM nonessential amino acids (Gibco; 11140050), 2.5 ml N2 supplement (Gibco; 17502048), 5 ml B27 supplement (Gibco; 17504044), 3 µM CHIR99021 (Sigma; SML1046), 1 µM PD 0325901 (Biomol; 13034-1), 10 ng/ml LIF (IMB Protein Production core facility). Cells were cultured at 37°C  with 5% CO2. Cell culture dishes were coated using 0.1% gelatin (Sigma; ES-006-B). The medium was exchanged every day and cells were passaged every second day.

extract_protocol:
For RNA extraction, cells were washed twice with ice cold 1x PBS and collected on ice. For total RNA isolation the RNeasy Plus Mini Kit (Qiagen) was used following the manufacturer’s instructions.
RNA-seq: All NGS library preps were performed with Illumina's Stranded mRNA Prep Ligation Kit following Stranded mRNA Prep Ligation Reference Guide (June 2020) (Document # 1000000124518 v00). For RNA-seq in HEK293T and C643 cells: Libraries were prepared with a starting amount of 350 ng for C643 samples and 1000 ng for HEK293T samples and amplified in 11 PCR cycles and 10 cycles respectively. For normalization, 2μl of a 1:300 dilution of ERCC spike-ins (Ambion) were added to the starting material for C643 samples and for HEK293T samples 2μl of a 1:100 dilution. Libraries were profiled in a DNA 1000 chip on a 2100 Bioanalyzer (Agilent technologies) and quantified using the Qubit dsDNA HS Assay Kit, in a Qubit 2.0 Fluorometer (Life technologies). All samples were pooled in equimolar ratio and sequenced on 1 NextSeq500 Highoutput FC, SR for 1x 80 cycles plus 10 cycles for the index read plus 1 dark cycle upfront Read 1. For RNAseq in male mESC cells upon 24 h of STM2457 treatment: Libraries were prepared with a starting amount of 1000 ng and amplified in 10 PCR cycles. For normalization, 2μl of a 1:100 dilution of ERCC spike-ins (Ambion) were added to the starting material. Libraries were profiled in a DNA 1000 chip on a 2100 Bioanalyzer (Agilent technologies) and quantified using the Qubit dsDNA HS Assay Kit, in a Qubit 2.0 Fluorometer (Life technologies). All samples were pooled in equimolar ratio and sequenced on 1 NextSeq500 Highoutput FC, SR for 1x 155 cycles plus 10 cycles for the index read and 1 dark cycle upfront Read 1. For RNAseq in RPE1 cells: Libraries were prepared with a starting amount of 1000 ng and amplified in 10 PCR cycles. For normalization, 2μl of a 1:100 dilution of ERCC spike-ins (Ambion) were added to the starting material. Libraries were profiled in a HS DNA chip on a 2100 Bioanalyzer (Agilent technologies) and quantified using the Qubit dsDNA HS Assay Kit, in a Qubit 2.0 Fluorometer (Life technologies). All samples were pooled in equimolar ratio and sequenced on 1 NextSeq500 Highoutput FC, SR for 1x 80 cycles plus 10 cycles for the index read and 1 dark cycle upfront Read 1. For RNAseq in male mESC upon STM2457 timecourse 3h-12 h: Libraries were prepared with a starting amount of 1000 ng and amplified in 10 cycles. For normalization, 2μl of a 1:100 dilution of ERCC spike-ins (Ambion) were added to the starting material. Libraries were profiled in a DNA 1000 chip on a 2100 Bioanalyzer (Agilent technologies) and quantified using the Qubit dsDNA HS Assay Kit, in a Qubit 2.0 Fluorometer (Life technologies). All samples were pooled in equimolar ratio and sequenced on 2 NextSeq500 Highoutput FC, SR for 1x 80 cycles plus 10 cycles for the index read plus 1 dark cycle upfront Read 1.  For RNAseq in female mESC single clones: Libraries were prepared with a starting amount of 500 ng and amplified in 10 PCR cycles. Two post PCR purification steps were performed to exclude residual primer and adapter dimers. Libraries were profiled in a DNA 1000 chip on a 2100 Bioanalyzer (Agilent technologies) and quantified using the Qubit dsDNA HS Assay Kit, in a Qubit 2.0 Fluorometer (Life technologies). All 12 samples were pooled in equimolar ratio and sequenced on 1 NextSeq500 Highoutput FC, SR for 1x 79 cycles plus 10 cycles for the index read and 2 dark cycles upfront Read 1.

treatment_protocol: Cells were cultured in respective medium supplemented with Mettl3 inhibitor STM2457 or DMSO as negative control for 3h and up to 24h.

tag: cell line

value: mESC

value: mouse embryonic stem cells (mESC)

tag: gender

value: male


title: mESC, STM2457, 9h, rep1


title: iN_46_dcre_d43_rep1

value: dcre


title: P0_WT_593_605_file3

description: single-cell RNA

data_processing:
Reads were counted and aggregated with 10x Genomics Cell Ranger (version 4.0.0)
Cells containing fewer than 200 or greater than 4000 features, or with mitochondrial genes making up over 15% of genes, were discarded using Seurat (version 4.0)
We used scArches (v0.3.5) with the scANVI model to calculate latent space of dimension ten, and to assign cell type annotations by mapping our data onto reference data using default parameters
Leiden clustering was performed on latent space (default parameters, k = 15). Two clusters with average annotation probability scores <80% were annotated based on marker gene expression; microglia with Tmem119 and Aif1, and blood cells with Hba-a1 and Hbb-bs. Blood cells were removed for downstream analysis.
Sub-populations of each cell type most affected by Myt1l mutation were identified using MELD (version 1.0) with parameters Beta=31 and KNN=5 which were determined using grid search
Differential expression was performed on these sub-populations using MAST (version 1.16.0) within Seurat’s FindMarkers function (logfc.threshold = 0, min.pct = 0.05, all other parameters default)
Genome_build: mm10
Supplementary_files_format_and_content: Tab-delimited file with MAST DE analysis (het vs wt)
Supplementary_files_format_and_content: Tab-delimited file with MAST DE analysis (hom vs wt)
Supplementary_files_format_and_content: Tab-delimited file with cell x gene matrix
Supplementary_files_format_and_content: Tab-delimited file with cell x metadata matrix

source_name: mouse cortex

extract_protocol:
Prefrontal cortices from P0 mice were dissected and dissociated into single cells and fixed in ice-cold 80% methanol and barcoded using the ECCITE-Seq scheme with an amine group at the 5’ end.
We pooled ~20.000 cells from two Myt1l (+/+), (+/-), and (-/-) animals, respectively. Each pool was loaded into an individual lane of the Chromium Controller (10x Genomics).
Chromium Single Cell 5' v1 reagent kit was used to process samples according to the manufacturer’s instructions, with modifications in the cDNA and library preparation steps suggested by the ECCITE-Seq protocol to generate the barcoded oligonucleotide libraries in parallel.
single-cell RNA-seq

treatment_protocol: Myt1l frameshift mutation was induced using CRISPR/Cas9 ribonucleoprotein (RNP) electroporation into mouse zygotes that caused a 7 bp deletion in Myt1l (ENSG00000186487) exon 6, resulting in a frameshift at amino acid (aa) 11 and terminating in a premature STOP at aa 78 in exon 9 (p.R11GfsX68). Myt1l mutant mice showed no nonsense-mediated decay of Myt1l transcript, but Western blot analysis showed expected depletion of full-length MYT1L, suggesting that this mouse model mimicked MYT1L deficiency reported in a patient with a premature STOP mutation at aa 75 (R75X).

tag: strain

tag: tissue

value: cortex

tag: age

tag: genotype

value: WT

tag: gem


title: Resolved Replicate 2, mRNA

city: South San Francisco

first: Joaquin

last: Pechuan jorge

state: CA

address: 1 dna way

department: Cancer Immunology

country: USA

institute: Genentech

description:
mRNA
LIB5424772_SAM24372269

data_processing:
Single-cell RNA-seq fastq files were processed with cellranger count (CellRanger 4.0.2 from 10x Genomics)using a custom reference based on the mouse genome GRCm38 with GENCODE35 gene  models  together  with  the  sequences  for  the  transgenes  employed  in  our  experiments  (tdTomatoand GFP). Similarly, TCR sequencing data was processed using cellranger vdj.  Analyses of the count matrices were conducted in R version 4.0.5 (2021-03-31) using Seurat39 (version 4.0.4). Only high quality cells were retained for the posterior analysis, more concretely, we kept the cells with more  than  300  hundred  genes  detected,  more  than  1000  unique  molecular  identifiers  (UMIs)  and  less than 10% mitochondrial reads.  To simplify our analyses, tdTomato positive cells (adoptive T cells) andGFP positive cells (KPP tumor cells) were separated and clustered independently.  The remaining cells were clustered to construct an atlas for stamp tumors.  In both cases, the clustering and identification of cell populations proceeded following Seurat’s SCTransform pipeline.  First, data was normalized usingSeurat::SCTransform() with cell cycle regression and batch correction.  Then,  the number of principal components retained for clustering was calculated using the talus plot40.  With these retained components, we then computed a UMAP embedding and the neighbors for posterior clustering. Several clustering  resolutions  were  calculated  and  a  directed  tree  was  constructed  reflecting  the  hier-archical  relationships  of  the  new  clusters  upon  increasing  the  resolution41.   A resolution was considered optimal if it did not break the hierarchical structure of said tree. The main clusters  were  identified  by  means  of  expression  markers  of  known  biology. Then,  for  each  major  cell cluster, a second run of clustering was performed by iterating the aforementioned pipeline.  Noticeably, this step allowed us to identify heterogeneity within our cell populations and further remove low quality cells. Markers for each cluster were identified using the Seurat::FindAllMarkers() method with default parameters, comparing all cells in a particular cluster to the rest of cells and accessing significantly dif-ferential gene expression using Wilcoxon’s rank sum test and Benjamini Hochberg correction for multiple testing.
Assembly: GRCm38
Supplementary files format and content: STAMPscAtlats.Rdata, contains the a seurat object with annotated clusters for the cells retained after quality control filtering.

source_name: Tumor

extract_protocol:
FOXN1-deficient nude mice with adoptively transferred tdTomato+ CD3+ T cells bearing KPP-EGFP STAMP microtumors were biopsied 8 days post tumor implantation using a 1 mm Miltex sterile disposable biopsy punch with plunger (Integra Biosciences, NH, USA). 3-6 pooled tissue biopsies were moved into a precooled 1.5 mL tube (Eppendorf, Germany) containing 300µL digestion cocktail consisting of Gibco RPMI 1640 (Thermo Fisher Scientific, MA, USA), 0.1% Gibco fetal calf serum (Thermo Fisher Scientific, MA, USA), 0.1mg/ml Liberase TM (Roche, Switzerland), 0.1mg/ml DNAse I (Roche, Switzerland), 32µM Gibco Actinomycin D (Thermo Fisher Scientific, MA, USA). Tissues were incubated for 30 min at 37°C and 950x RPM on a Thermoblock (Eppendorf, Germany) and mechanically dissociated every 10 min with a pipette. To quench the digestion, the cell suspension was filtered through a 40 µm mesh into pre-cooled FACS filter tube containing quenching buffer of Gibco fetal calf serum (Thermo Fisher Scientific, MA, USA) with 32 µM Gibco Actinomycin D (Thermo Fisher Scientific, MA, USA). The cell suspension was centrifuged at 350xg for 8 min and resuspended in 400 µL Gibco RPMI 1640 (Thermo Fisher Scientific, MA, USA) with 5 µM CalceinBlue (Invitrogen, CA, USA) and a 1:200 dilution of Molecular Probes Fixable Live/Dead Near-IR Dead Cell Stain Kit (Thermo Fisher Scientific, MA, USA). FACS was performed to isolate Calcein Blue-positive and L/D NearIR-negative cells into a precooled 1.5ml Eppendorf tube containing 750µL collection buffer consisting of Gibco RPMI 1640 (Thermo Fisher Scientific, MA, USA), 10% Gibco fetal calf serum (Thermo Fisher Scientific, MA, USA), 32 µM Gibco Actinomycin D (Thermo Fisher Scientific, MA, USA). The cell number and viability were determined on a Vi-Cell XR cell viability analyzer (Beckman Coulter, CA, USA) and scRNAseq library preparation was performed using dual single cell mouse kit 5’/TCR according to the manufacturer’s instructions (10x Genomics, CA, USA).
Gene Expression and paired TCR sequencing for tumors coming from the different immune phenotypes (inflamed, excluded or desert) plus some reference types (resolved, empty pores and tumors coming from a mouse without adoptively transferred Tcells.

value: Tumor

value: KPP-EGFP

tag: primary tissue

value: Skin

value: STAMP Tumor Resolved Phenotype


title: Resolved Replicate 2, TCRseq

description:
mRNA, TCRseq
LIB5425676_SAM24372269


title: P0_WT_593_605_file2

description: single-cell RNA


title: Mrps5 fl/fl mouse heart tissue, control, 12 weeks after tamoxifen injection, rep 2

city: Hang Zhou

first: Feng

last: Gao

state: Zhejiang

address: Kaixuan road 268

department: Department of Cardiology

country: China

institute: Second Afliated Hospital, Zhejiang University School of Medicine

data_processing:
The clustering of the index-coded samples was performed on a cBot Cluster Generation System using TruSeq PE Cluster Kit v3-cBot-HS (Illumia) according to the manufacturer’s instructions. After cluster generation, the library preparations were sequenced on an Illumina Hiseq platform and 125 bp/150 bp paired-end reads were generated.
Raw data (raw reads) of fastq format were firstly processed through in-house perl scripts. In this step, clean data (clean reads) were obtained by removing reads containing adapter, reads containing ploy-N and low quality reads from raw data. At the same time, Q20, Q30 and GC content the clean data were calculated. All the downstream analyses were based on the clean data with high quality.
Reference genome and gene model annotation files were downloaded from genome website directly. Index of the reference genome was built using Hisat2 v2.0.5 and paired-end clean reads were aligned to the reference genome using Hisat2 v2.0.5. We selected Hisat2 as the mapping tool for that Hisat2 can generate a database of splice junctions based on the gene model annotation file and thus a better mapping result than other non-splice mapping tools.
featureCounts v1.5.0-p3 was used to count the reads numbers mapped to each gene. And then FPKM of each gene was calculated based on the length of the gene and reads count mapped to this gene. FPKM, expected number of Fragments Per Kilobase of transcript sequence per Millions base pairs sequenced, considers the effect of sequencing depth and gene length for the reads count at the same time, and is currently the most commonly used method for estimating gene expression levels.
Differential expression analysis of two conditions/groups (two biological replicates per condition) was performed using the DESeq2 R package (1.16.1). DESeq2 provide statistical routines for determining differential expression in digital gene expression data using a model based on the negative binomial distribution. The resulting P-values were adjusted using the Benjamini and Hochberg’s approach for controlling the false discovery rate . Genes with an adjusted P-value <0.05 found by DESeq2 were assigned as differentially expressed.
Gene Ontology (GO) enrichment analysis of differentially expressed genes was implemented by the clusterProfiler R package, in which gene length bias wascorrected. GO terms with corrected Pvalue less than 0.05 were considered significantly enriched by differential expressed genes. KEGG is a database resource for understanding high-level functions and utilities of the biological system, such as the cell, the organism and the ecosystem, from molecular-level information, especially large-scale molecular datasets generated by genome sequencing and other high-through put experimental technologies (http://www.genome.jp/kegg/). We used clusterProfiler R package to test the statistical enrichment of differential expression genes in KEGG pathways.
Assembly: mm9
Supplementary files format and content: tab-delimited text files include FPKM values for each Sample

source_name: Heart

growth_protocol: Mrps5 fl/fl mouse and Mrps5 fl/fl; MCM mouse were generated and grown up to 8 weeks old in SPF animal labratory

extract_protocol:
RNA was harvested using Trizol (Life). RNA degradation and contamination was monitored on 1% agarose gels. RNA purity was checked using the NanoPhotometer® spectrophotometer (IMPLEN, CA, USA) . RNA concentration was measured using Qubit® RNA Assay Kit in Qubit®2.0 Flurometer (Life Technologies, CA, USA). RNA integrity was assessed using the RNA Nano 6000 Assay Kit of the Bioanalyzer 2100 system (Agilent Technologies, CA, USA).
A total amount of 3 μg RNA per sample was used as input material for the RNA sample preparations. Sequencing 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. Briefly, mRNA was purified from total RNA using poly-T oligo-attached magnetic beads. Fragmentation was carried out using divalent cations under elevated temperature in NEBNext First Strand Synthesis Reaction Buffer(5X). First strand cDNA was synthesized using random hexamer primer and M-MuLV Reverse Transcriptase(RNase H-). Second strand cDNA synthesis was subsequently  performed using DNA Polymerase I and RNase H. Remaining overhangs were converted into blunt ends via exonuclease/polymerase activities. After adenylation of 3’ ends of DNA fragments, NEBNext Adaptor with hairpin loop structure were ligated to prepare for hybridization. In order to select cDNA fragments of preferentially 250~300 bp in length, the library fragments were purified with AMPure XP system (Beckman Coulter, Beverly, USA). Then 3 μl USER Enzyme (NEB, USA) was used with size-selected, adaptor-ligated cDNA at 37°C for 15 min followed by 5 min at 95 °C before PCR. Then PCR was performed with Phusion High-Fidelity DNA polymerase, Universal PCR primers and Index (X) Primer. At last, PCR products were purified (AMPure XP system) and library quality was assessed on the Agilent Bioanalyzer 2100 system.

treatment_protocol: 8 weeks old Mrps5 fl/fl mouse and Mrps5 fl/fl; MCM mouse were treated with tamoxifen through intraperitoneal injection at the dosage of 45 mg/kg/day for 5 continuous days to activate Cre recombinase and delete Mrps5 gene in genome.

value: Mrps5 fl/fl

tag: time

value: 12 weeks after tamoxifen injection


title: mESC, STM2457, 6h, rep1


title: mESC, STM2457, 6h, rep2


title: Resolved Replicate 1, mRNA

description:
mRNA
LIB5419349_SAM24363084


title: P0_WT_593_605_file1

description: single-cell RNA


title: iN_46_dcre_d7_rep1


title: iN_46_dcre_d7_rep2


title: P0_HOM_592_604_file4

description: single-cell RNA

value: HOM


title: Resolved Replicate 1, TCRseq

description:
mRNA, TCRseq
LIB5423216_SAM24363084


title: mESC, DMSO, 6h, rep4

value: DMSO


title: P0_HOM_592_604_file3

description: single-cell RNA


title: Inflamed Replicate 2, mRNA

description:
mRNA
LIB5424770_SAM24372267

value: STAMP Tumor Inflamed Phenotype


title: Mrps5 fl/fl mouse heart tissue, control, 12 weeks after tamoxifen injection, rep 1

value: Heart

value: Tamoxifen injection


title: mESC, DMSO, 6h, rep3

value: mESC


title: iN_33_cre_d43_rep1

value: neuron

value: cre

value: clone 2


title: iN_33_cre_d43_rep2


title: mESC, STM2457, 6h, rep3

value: mouse embryonic stem cells (mESC)

value: male


title: iN_46_cre_d7_rep1


title: mESC, DMSO, 6h, rep2


title: iN_33_dcre_d43_rep2

value: dcre


title: mESC, DMSO, 6h, rep1


title: Inflamed Replicate 1, TCRseq

description:
mRNA, TCRseq
LIB5423214_SAM24363082

value: Tumor

value: KPP-EGFP


title: Inflamed Replicate 1, mRNA

description:
mRNA
LIB5419347_SAM24363082

value: Skin


title: P0_HOM_592_604_file2

description: single-cell RNA

value: cortex


title: P0_HET_591_607_file4

description: single-cell RNA

value: HET


title: P0_HOM_592_604_file1

description: single-cell RNA

value: HOM


title: iN_33_cre_d7_rep2


title: P0_HET_591_607_file3

description: single-cell RNA


title: Excluded Replicate 2, TCRseq

description:
mRNA, TCRseq
LIB5425675_SAM24372268

value: STAMP Tumor Excluded Phenotype


title: Excluded Replicate 2, mRNA

description:
mRNA
LIB5424771_SAM24372268


title: iN_33_cre_d7_rep1


title: mESC, STM2457, 3h, rep4


title: P0_HET_591_607_file2

description: single-cell RNA


title: mESC, STM2457, 3h, rep3


title: mESC, STM2457, 3h, rep2


title: Inflamed Replicate 2, TCRseq

description:
mRNA, TCRseq
LIB5425674_SAM24372267

value: STAMP Tumor Inflamed Phenotype


title: P0_HET_591_607_file1

description: single-cell RNA


title: mESC, STM2457, 3h, rep1


title: iN_33_dcre_d7_rep2


title: E18.5_WT_735_rep1

data_processing:
Sequenced reads were trimmed for adaptor sequence and masked for low-complexity or low-quality sequence
Filtered reads were mapped to mm10 with STAR to generate raw counts files
Differential expression and normalised counts were determined using DESeq2 (R package version 1.28.1) with sequencing batch ID and sex as covariates
Genome_build: mm10
Supplementary_files_format_and_content: Tab-delimited text file with raw counts for all samples
Supplementary_files_format_and_content: Tab-delimited text file with DESeq2 output for all conditions (sample-collapsed)
Supplementary_files_format_and_content: Tab-delimited text file with normalised counts for all samples

extract_protocol:
The cortex of E18.5 or the prefrontal cortex of P0, P22, and adult (3-month-old) mice was dissected and placed into TRIzol (Invitrogen) followed by RNA harvest using RNA Miniprep kit (Zymo Research).
RNA libraries were prepared for sequencing using standard Illumina protocols

value: WT

tag: Sex


title: iN_33_dcre_d43_rep1


title: iN_33_dcre_d7_rep1


title: Excluded Replicate 1, mRNA

description:
mRNA
LIB5419348_SAM24363083


title: E18.5_WT_735_rep2


title: mESC, DMSO, 3h, rep4

value: DMSO


title: Desert Replicate 2, mRNA

description:
mRNA
LIB5424773_SAM24372270

value: STAMP Tumor Desert Phenotype


title: iN_31_cre_d43_rep1

value: neuron


title: mESC, DMSO, 3h, rep3

value: mESC


title: iN_34_cre_d43_rep2

value: cre

value: clone 1


title: iN_31_cre_d43_rep2

value: clone 2


title: E18.5_HET_733_rep2

value: male


title: Desert Replicate 2, TCRseq

description:
mRNA, TCRseq
LIB5425677_SAM24372270

value: Tumor


title: E18.5_HET_733_rep1


title: iN_31_dcre_d43_rep2

value: dcre


title: mESC, DMSO, 3h, rep2

value: mouse embryonic stem cells (mESC)


title: iN_31_dcre_d43_rep1


title: Excluded Replicate 1, TCRseq

description:
mRNA, TCRseq
LIB5423215_SAM24363083

value: KPP-EGFP

value: Skin


title: Desert Replicate 1, mRNA

description:
mRNA
LIB5419350_SAM24363085


title: iN_31_cre_d7_rep2


title: Desert Replicate 1, TCRseq

description:
mRNA, TCRseq
LIB5423217_SAM24363085


title: E18.5_WT_732_rep1

value: cortex


title: E18.5_WT_732_rep2


title: mESC, STM2457, rep4


title: iN_31_cre_d7_rep1


title: mESC, STM2457 rep3


title: mESC, STM2457, rep2


title: iN_31_dcre_d7_rep2


title: No Tcells, mRNA

description:
mRNA
LIB5424774_SAM24372271

value: STAMP Tumor with no T Cells


title: E18.5_HOM_730_rep1

value: HOM


title: E18.5_HOM_730_rep2


title: Empty Pore, TCRseq

description:
mRNA, TCRseq
LIB5425678_SAM24372272

source_name: Skin

value: STAMP Pore


title: Excluded [SAM24376860]

data_processing:
Each RNA-sequencing experiment was analyzed using HTSeqGenie pipeline (Pau and Reeder, 2012) inBioConductor34  as  follows:  first,  reads  with  low  nucleotide  qualities  (70% of bases with quality < 23) or matches to rRNA and adapter sequences were removed.  The remaining reads were aligned to the mouse reference genome GRCm38.p5 using GSNAP (Wu & Nacu 2010, Wu & Wu 2016) version ‘2013-10-10-v2’,  allowing maximum of two mismatches per 75 base sequence (parameters:  ‘-M2 -n 10 -B 2 -i 1 -N 1 -w 200000 -E 1 –pairmax-rna=200000 –clip-overlap’).  Transcript annotation was based  on  the  Gencode  genes  database35. To quantify  gene  expression  levels,  the number of reads mapping unambiguously to the exons of each gene was calculated. he count matrix was filtered  to remove lowly expressed genes by keeping the features with at least 0.2 counts per million (cpm)  in more than the minimum number of samples of the experimental group given the design factor  being analyzed. The resulting filtered count matrix was then log transformed and TMM normalized  with edgeR::cpm(log=T) and edgeR::calcNormFactors(method=”TMM”) in order to perform ex-  ploratory analyses.
Assembly: GRCm38
Supplementary files format and content: Day13OnlyImmuneTMMLogCPM.csv TMM normalized logCPM matrix, rows are genes and columns are samples by SAMID

extract_protocol: Each tissue biopsy was transferred to a separate 1.5 mL tube (Eppendorf, Germany) containing 0.25 mL Invitrogen TRIzol Reagent (Thermo Fisher Scientific, MA, USA). Biopsies were incubated in TRIzol for 5 min with intermittent vortexing. 50 µL chloroform (MilliporeSigma, MA, USA) was added to the homogenate, vortexed for 20 sec, and incubated at 20-25°C for 2-3 min. To accelerate phase separation, samples were centrifuged at 10,000xg for 18 min at 4°C. The aqueous (top) phase was removed by aspiration and transferred to a clean 1.5 mL tube (Eppendorf, Germany). A volume of 100% RNAse-free ethanol (MilliporeSigma, MA, USA). equal to the volume of the aqueous layer was added, and the RNA was further isolated using a RNeasy Micro Kit (Qiagen, Germany). Alternatively, an individual tumor biopsy was immersed in RNA later and the RNA further extracted using a RNeasy Mini Kit (Qiagen, Germany).

tag: phenotype

value: Excluded


title: mESC, STM2457, rep1


title: mESC, DMSO, rep4

value: DMSO


title: mESC, DMSO, 3h, rep1


title: Empty Pore, mRNA

description:
mRNA
LIB5424775_SAM24372272


title: Excluded [SAM24376859]


title: 1-D-G-Desert BULK [SAM24366208]

data_processing:
Each RNA-sequencing experiment was analyzed using HTSeqGenie pipeline (Pau and Reeder, 2012) inBioConductor34  as  follows:  first,  reads  with  low  nucleotide  qualities  (70% of bases with quality < 23) or matches to rRNA and adapter sequences were removed.  The remaining reads were aligned to the mouse reference genome GRCm38.p5 using GSNAP (Wu & Nacu 2010, Wu & Wu 2016) version ‘2013-10-10-v2’,  allowing maximum of two mismatches per 75 base sequence (parameters:  ‘-M2 -n 10 -B 2 -i 1 -N 1 -w 200000 -E 1 –pairmax-rna=200000 –clip-overlap’).  Transcript annotation was based  on  the  Gencode  genes  database35. To quantify  gene  expression  levels,  the number of reads mapping unambiguously to the exons of each gene was calculated. he count matrix was filtered to remove lowly expressed genes by keeping the features with at least 0.2 counts per million (cpm) in more than the minimum number of samples of the experimental group given the design factor being analyzed. The resulting filtered count matrix was then log transformed and TMM normalized with edgeR::cpm(log=T) and edgeR::calcNormFactors(method=”TMM”) in order to perform exploratory analyses.
Assembly: GRCm38
Supplementary files format and content: D8OnlyImmuneTMMLogCPM.csv rows are genes and columns are samples by SAMID


title: E18.5_HET_729_rep1

value: HET

value: female


title: E18.5_HET_729_rep2


title: Excluded [SAM24376858]


title: 1-D-F-Desert BULK [SAM24366207]


title: E18.5_HOM_728_rep2


title: mESC, DMSO, rep3

value: mESC


title: mESC, DMSO, rep1


title: mESC, DMSO, rep2


title: 1-D-D-Desert BULK [SAM24366205]


title: P0_HET_675_rep1


title: Excluded [SAM24376857]


title: female mESC, single clone40

description: female

library_source: genomic

data_processing:
Quality control: FastQC (v0.11.8)
Trimming: Cutadapt (v2.4) with non-default parameters: cutadapt -a file:adapters.fa --minimum-length 20 (adapter.fa: >Illumina CTGTCTCTTATACACATCT >NEBNext GATCGGAAGAGCACACGT)
Mapping: Bowtie2 (v2.3.4.3) with non-default parameters: bowtie2 -q --phred33 -N 0 -L 31 -i S,1,0.50 -D 20 -R 3 -p 4 --end-to-end
Filter/remove low quality or multimapping hits: Samtools (v1.9) with parameters: samtools view -F 4 -q 3 -bhu
Remove duplicates: Picard (v2.20.3) with non-default parameters: java -Xmx5000m -jar picard.jar MarkDuplicates REMOVE_DUPLICATES=TRUE ASSUME_SORTED=TRUE
Assembly: GRCm38.p6 (Gencode release M23)
Supplementary files format and content: Reads were counted in equal sized bins (100 kb) using the countOverlaps() function from the IRanges package (v2.28.0) in R. Counts are given in bedGraph file format.
Library strategy: DNA-seq

molecule: genomic DNA

growth_protocol: Female mouse embryonic stem cells (mESC) were taken from a previous publication (PMID: 24506884). Single clones were picked and cells were cultured in 2i/LIF medium. Briefly, 500 ml medium was generated by adding 235 ml of each DMEM/F12 and neurobasal (Gibco; 21331020, 21103049), 7.5 ml BSA solution (7.5%, ThermoFisher; 11500496), 5 ml penicillin-streptomycin (Gibco; 15140-122), 2 mM L-Glutamin (ThermoSci; 25030024), 100 µM β-mercaptoethanol (Gibco; 21985023), 5 ml mM nonessential amino acids (Gibco; 11140050), 2.5 ml N2 supplement (Gibco; 17502048), 5 ml B27 supplement (Gibco; 17504044), 3 µM CHIR99021 (Sigma; SML1046), 1 µM PD 0325901 (Biomol; 13034-1), 10 ng/ml LIF (IMB Protein Production core facility). Cells were cultured at 37°C  with 5% CO2. Cell culture dishes were coated using 0.1% gelatin (Sigma; ES-006-B). The medium was exchanged every day and cells were passaged every second day.

extract_protocol:
Cells were washed twice with ice cold 1x PBS and collected in ice. For DNA isolation the PureLink Genomic DNA MINI Kit (Invitrogen) was used following the manufacturer’s instructions.
gDNA was sheared with a Covaris E220 focused ultrasonicator (Parameters: Peak Incident Power (W)=18, Duty Cycle=20%, Cycles per burst=50, Temperature=20°C, Water Level=6, Treatment Time=80sec). NGS library prep was performed using ½ rxn of NEBNext Ultra II DNA Library Prep Kit for Illumina Version 6.0, 3/20. Libraries were prepared with a starting amount of 20ng fragmented DNA and were amplified in 6 PCR cycles. Libraries were profiled in a High Sensitivity DNA on a 2100 Bioanalyzer (Agilent technologies) and quantified using the Qubit dsDNA HS Assay Kit, in a Qubit 2.0 Fluorometer (Life technologies). All 20 samples were pooled in equimolar ratio on a NextSeq500 Midoutput FC, SR for 1x 159 cycles plus 7 cycles for the index read.

value: female mouse embryonic stem cells (mESC)


title: 1-D-C-Desert BULK [SAM24366204]


title: Inflamed [SAM24376855]

value: Inflamed


title: iN_31_dcre_d7_rep1

value: neuron

value: clone 2


title: Inflamed [SAM24376854]

value: Tumor


title: P0_HET_675_rep2

value: male


title: P0_WT_674_rep1

value: WT


title: 1-D-B-Desert BULK [SAM24366203]

value: KPP-EGFP

value: Skin


title: Inflamed [SAM24376853]


title: female mESC, single clone38

description: female


title: 1-D-A-Desert BULK [SAM24366202]


title: P0_WT_674_rep2

value: cortex


title: 1-E-G-Excluded BULK [SAM24366194]


title: 1-D-E-Desert BULK [SAM24366206]


title: P0_HET_672_rep2


title: female mESC, single clone37

description: female


title: Inflamed [SAM24376851]


title: 1-E-F-Excluded BULK [SAM24366193]


title: Inflamed [SAM24376850]


title: P0_HET_672_rep1


title: 1-E-E-Excluded BULK [SAM24366192]


title: female mESC, single clone35

description: female


title: E18.5_HOM_728_rep1

value: HOM


title: Inflamed [SAM24376849]


title: female mESC, single clone39

description: female


title: P0_HOM_671_rep2


title: 1-E-D-Excluded BULK [SAM24366191]


title: P0_HOM_671_rep1


title: Desert [SAM24376841]

value: Desert


title: female mESC, single clone34

description: female


title: 1-E-C-Excluded BULK [SAM24366190]


title: P0_HOM_670_rep1


title: female mESC, single clone33

description: female


title: female mESC, single clone32

description: female


title: female mESC, single clone36

description: female


title: P0_HOM_670_rep2


title: 1-E-A-Excluded BULK [SAM24366188]


title: female mESC, single clone31

description: female


title: Desert [SAM24376840]


title: P0_HET_669_rep2


title: Desert [SAM24376838]


title: female mESC, single clone10

description: female


title: 1-H-G-Inflamed BULK [SAM24366187]


title: P0_HET_669_rep1


title: 1-H-F-Inflamed BULK [SAM24366186]


title: Desert [SAM24376837]


title: female mESC, single clone9

description: female


title: 1-H-E-Inflamed BULK [SAM24366185]


title: Adult_HET_1495_rep1

value: adult


title: 1-H-D-Inflamed BULK [SAM24366184]


title: 1-E-B-Excluded BULK [SAM24366189]


title: Excluded [SAM24376828]

value: Excluded


title: 1-H-C-Inflamed BULK [SAM24366183]


title: Mrps5 fl/fl; cTNT-Cre mouse heart tissue, cKO, embryonic 12 days, rep 3

data_processing:
Raw data(raw reads) of fastq format were firstly processed through in-house perl scripts. In this step, clean data(clean reads) were obtained by removing reads containing adapter, reads on containing ploy-N and low quality reads from raw data. At the same time, Q20, Q30 and GC content of the clean data were calculated. All the down stream analyses were based on the clean data with high quality.  Reference genome and gene model annotation files were downloaded from genome website directly. Index of the reference genome was built using bowtie2 v2.2.8 and paired-end clean reads were aligned to the reference genome using HISAT2(Langmead, B.et al) v2.0.4. HISAT2 was run with ‘--rna-strandness RF’, other parameters were set as default. The mapped reads of each sample were assembled by StringTie (v1.3.1) (Mihaela Pertea.et al. 2016) in a reference-based approach. StringTie uses a novel network flow algorithm as well as an optional de novo assembly step to assemble and quantitate full- length transcripts representing multiple splice variants for each gene locus.
CNCI (Coding-Non-Coding-Index) (v2) profiles adjoining nucleotide triplets to effectively distinguish protein-coding and non-coding sequences independent of known annotations (Sun et al. 2013). We use CNCI with default parameters.  CPC (Coding Potential Calculator) (0.9-r2) mainly through assess the extent and quality of the ORF in a transcript and search the sequences with known protein sequence database to clarify the coding and non-coding transcripts (Kong et al. 2007). We used the NCBI eukaryotes' protein database and set the e-value ‘1e-10’ in our analysis. We translated each transcript in all three possible frames and used Pfam Scan (v1.3) to identify occurrence of any of the known protein family domains documented in the Pfam database (release 27; used both Pfam A and Pfam B)(Punta, et al. 2012). Any transcript with a Pfam hit would be excluded in following steps. Pfam searches use default parameters of -E 0.001 --domE 0.001 (Bateman, et al. 2002). PhyloCSF (phylogenetic codon substitution frequency) (v20121028) examines evolutionary signatures characteristic to alignments of conserved coding regions, such as the high frequencies of synonymous codon substitutions and conservative amino acid substitutions, and the low frequencies of other missense and non-sense substitutions to distinguish protein-coding and non-coding transcripts (Lin et al. 2011). We build multi-species genome sequence alignments and run phyloCSF with default parameters. Transcripts predicted with coding potential by either/all of the four tools above were filtered out, and those without coding potential were our candidate set of lncRNAs.
Phast (v1.3) is a software package contains much of statistical programs, most used in phylogenetic analysis (Siepel, et al. 2005), and phastCons is a conservation scoring and identificating program of conserved elements. We used phyloFit to compute phylogenetic models for conserved and non-conserved regions among species and then gave the model and HMM transition parameters to phyloP to compute a set of conservation scores of lncRNA and coding genes.
Cis role of target gene prediction: Cis role is lncRNA acting on neighboring target genes. We searched coding genes 10k/100k upstream and downstream of lncRNA and then analyzed their function next. Trans role of target gene prediction: Trans role is lncRNA to identify each other by the expression level. While there were no more than 25 samples, we calculated the expressed correlation between lncRNAs and coding genes with custom scripts; otherwise, we clustered the genes from different samples with WGCNA (Langfelder et al, 2008) to search common expression modules and then analyzed their function through functional enrichment analysis.
Cuffdiff (v2.1.1) was used to calculate FPKMs of both lncRNAs and coding genes in each sample (Trapnell, C. et al. 2010). Gene FPKMs were computed by summing the FPKMs of transcripts in each gene group. FPKM means fragments per kilo-base of exon per million fragments mapped, calculated based on the length of the fragments and reads count mapped to this fragment.  The Ballgown suite includes functions for interactive exploration of the transcriptome assembly, visualization of transcript structures and feature-specific abundances for each locus, and post-hoc annotation of assembled features to annotated features(Alyssa C. Frazee et al.2014). Transcripts with an P-adjust <0.05 were assigned as differentially expressed. Cuffdiff provides statistical routines for determining differential expression in digital transcript or gene expression data using a model based on the negative binomial distribution (Trapnell, C. et al. 2010). Transcripts with an P-adjust <0.05 were assigned as differentially expressed.
Gene Ontology (GO) enrichment analysis of differentially expressed genes or lncRNA target genes were implemented by the GOseq R package, in which gene length bias was corrected(Young, M. D.et al.2010). GO terms with corrected Pvalue less than 0.05 were considered significantly enriched by differential expressed genes. KEGG is a database resource for understanding high-level functions and utilities of the biological system(Kanehisa, M.et al.2008), such as the cell, the organism and the ecosystem, from molecular-level information, especially large-scale molecular datasets generated by genome sequencing and other high-throughput experimental technologies (http://www.genome.jp/kegg/). We used KOBAS software to test the statistical enrichment of differential expression genes or lncRNA target genes in KEGG pathways(Mao, X.et al.1995).
Assembly: mm9
Supplementary files format and content: tab-delimited text files include FPKM values for each Sample

growth_protocol: To gain insight into the function of Mrps5 in cardiac development, we crossed cTNT-Cre mice with Mrps5 fl/fl mice, then used the obtained Mrps5 fl/+; cTNT-Cre mice cross with Mrps5 fl/fl mice again to obtain Mrps5 fl/fl and Mrps5 fl/fl; cTNT-Cre mouse embyros

extract_protocol:
RNA was harvested using Trizol (Life). RNA degradation and contamination was monitored on 1% agarose gels. RNA purity was checked using the NanoPhotometer® spectrophotometer  (IMPLEN, CA, USA). RNA concentration was measured using Qubit® RNA Assay Kit in Qubit® 2.0 Flurometer (Life Technologies, CA, USA). RNA integrity was assessed using the RNA Nano 6000 Assay Kit of the Bioanalyzer 2100 system (Agilent Technologies, CA, USA).
A total amount of 3 μg RNA per sample was used as input material for the RNA sample preparations. Firstly, ribosomal RNA was removed by Epicentre Ribo-zeroTM rRNA Removal Kit (Epicentre, USA), and rRNA free residue was cleaned up by ethanol precipitation. Subsequently, sequencing libraries were generated using the rRNA- depleted RNA by NEBNext® UltraTM Directional RNA Library Prep Kit for Illumina® (NEB, USA) following manufacturer’s recommendations. Briefly, fragmentation was carried out using divalent cations under elevated temperature in NEBNext First Strand Synthesis Reaction Buffer(5X). First strand cDNA was synthesized using random hexamer primer and M-MuLV Reverse Transcriptase(RNaseH-). Second strand cDNA synthesis was subsequently performed using DNA Polymerase I and RNase H. In the reaction buffer, dNTPs with dTTP were replaced by dUTP. Remaining overhangs were converted into blunt ends via exonuclease/polymerase activities. After adenylation of 3’ ends of DNA fragments, NEBNext Adaptor with hairpin loop structure were ligated to prepare for hybridization. In order to select cDNA fragments of preferentially 150~200 bp in length, the library fragments were purified with AMPure XP system (Beckman Coulter, Beverly, USA). Then 3 μl USER Enzyme (NEB, USA) was used with size-selected, adaptor-ligated cDNA at 37° C for 15 min followed by 5 min at 95°C before PCR. Then PCR was performed with Phusion High-Fidelity DNA polymerase, Universal PCR primers and Index (X) Primer. At last, products were purified (AMPure XP system) and library quality was assessed on the Agilent Bioanalyzer 2100 system. The clustering of the index-coded samples was performed on a cBot Cluster Generation System using TruSeq PE Cluster Kit v3-cBot-HS (Illumia) according to the manufacturer’s instructions. After cluster generation, the libraries were sequenced on an Illumina Hiseq 4000 platform and 150 bp paired-end reads were generated.

treatment_protocol: To further assess the cellular mechanism underlying myocardial hypoplasia of the Mrps5 mutant hearts, we performed the transcriptome sequencing on the Mrps5 mutants hearts and its control littermates, hearts were collected at embryonic day 12, prior to the onset of embryos lethality. Each group has 3 biological replicates, control group indicates as Ctrl_1, Ctrl_2, Ctrl_3, and the cKO group indicates as cKO_1, cKO_2, cKO_3.

value: Heart

value: Mrps5 fl/fl; cTNT-Cre

tag: development stage

value: Embryo

tag: time point

value: Embryonic 12 days


title: female mESC, single clone7

description: female

tag: cell type

value: female mouse embryonic stem cells (mESC)


title: Mrps5 fl/fl; cTNT-Cre mouse heart tissue, cKO, embryonic 12 days, rep 2


title: Desert [SAM24376839]


title: female mESC, single clone6

description: female


title: Adult_HET_1496_rep2

value: HET


title: Adult_HET_1496_rep1


title: 1-H-B-Inflamed BULK [SAM24366182]


title: Adult_HET_1492_rep2

value: female


title: female mESC, single clone4

description: female


title: female mESC, single clone5

description: female


title: Mrps5 fl/fl; cTNT-Cre mouse heart tissue, cKO, embryonic 12 days, rep 1


title: Adult_HET_1492_rep1


title: female mESC, single clone3

description: female


title: female mESC, single clone2

description: female


title: Mrps5 fl/fl mouse heart tissue, ctrl, embryonic 12 days, rep 3

value: Mrps5 fl/fl


title: Adult_HET_1495_rep2

value: male


title: 1-H-A-Inflamed BULK [SAM24366181]

value: Tumor


title: Excluded [SAM24376829]

value: KPP-EGFP


title: Excluded [SAM24376856]

value: Skin


title: Skin_Day8 [SAM24405550]

data_processing:
Each RNA-sequencing experiment was analyzed using HTSeqGenie pipeline (Pau and Reeder, 2012) inBioConductor34  as  follows:  first,  reads  with  low  nucleotide  qualities  (70% of bases with quality < 23) or matches to rRNA and adapter sequences were removed.  The remaining reads were aligned to the mouse reference genome GRCm38.p5 using GSNAP (Wu & Nacu 2010, Wu & Wu 2016) version ‘2013-10-10-v2’,  allowing maximum of two mismatches per 75 base sequence (parameters:  ‘-M2 -n 10 -B 2 -i 1 -N 1 -w 200000 -E 1 –pairmax-rna=200000 –clip-overlap’).  Transcript annotation was based  on  the  Gencode  genes  database35. To quantify  gene  expression  levels,  the number of reads mapping unambiguously to the exons of each gene was calculated.
Assembly: GRCm38

tag: day


title: female mESC, single clone8

description: female


title: Skin_Day8 [SAM24405549]


title: E18.5_HET_710_rep2

value: cortex


title: Skin_Day8 [SAM24405548]


title: RNA_siNUDT5 rep3

city: BOSTON

first: HONGYUN

last: QI

address: 450 Brookline Ave

institute: RADIATION ONCOLOGY

data_processing:
For RNA-seq, Sequenced reads were aligned to the UCSC hg19 reference genome assembly and gene counts were quantified using STAR (v2.7.3a). Differential gene expression testing was performed by DESeq2 (v1.22.1). RNAseq analysis was performed using the VIPER snakemake pipeline.
Genome_build: hg 19
Supplementary_files_format_and_content: bigwig

growth_protocol: U2OS cells were cultured in Dulbecco’s modified Eagle’s medium containing 10% fetal bovine serum.

extract_protocol:
For RNA-seq, RNA was extracted using RNeasy Mini kit (Qiagen).
For RNA-seq, Libraries were prepared using Roche Kapa mRNA HyperPrep strand specific sample preparation kits from 200ng of purified total RNA according to the manufacturer’s protocol on a Beckman Coulter Biomek i7.

treatment_protocol: For RNA-seq, U2OS cells were transfected with siRNAs and recovered for 48 hours before RNA extraction.

tag: si rna

value: transfected with siRNA targeting NUDT5

tag: treatment

value: untreated


title: E18.5_WT_709_rep2

value: WT


title: Skin_Day8 [SAM24405547]

tag: cell line


title: 1867_G1E-ER4_+E2_24_hours_PRO-seq_rep2

city: Philadelphia

first: Marit

middle: W.

last: Vermunt

state: PA

address: 3615 Civic Center Blvd

department: Hematology

institute: Children's Hospital of Phildelphia

data_processing:
Library strategy: PRO-seq
Read pairs originated from short RNA fragments were merged with FLASH2
PCR duplicates were removed using 6-base UMI barcodes, which were trimmed from the reads afterwards
Both merged single end reads and unmerged paired end reads were aligned to reference genome (mm9) using BWA-MEM (Li and Durbin, 2010)
Primary alignments with no INDELs and mapq score of at lease 20 were reported
Genome_build: mm9
Supplementary_files_format_and_content: bigwig - pileup of aligned PRO-seq read pairs
Supplementary_files_format_and_content: text - number of read pairs mapped to dREG peaks and gene bodies

growth_protocol: G1E and G1E-ER4 cells were grown in IMDM media (Corning, 10-016-CV) with 15% fetal calf serum (Gemini, lot A08H00K), 2% penicillin/streptomycin (Gibco, 15140-122), 140 μm 1-thioglycerol (Sigma, M6145), 2 units/ml epoetin (Amgen) and conditioned medium from a kit ligand-producing CHO cell line. Cells were grown at a cell density of 100,000 to 1,000,000 cells/ml.

extract_protocol: PRO-seq protocol was performed as previously described: Mahat et al., 2016. Base-pair-resolution genome-wide mapping of active RNA polymerases using precision nuclear run-on (PRO-seq).

treatment_protocol: Erythroid maturation experiments were performed by plating cells at a cell density of ~200,000 cells/ml in fresh G1E medium containing 100 nm estradiol (Sigma, E2758) for durations indicated for each experiment.

value: Estradiol (E2) for 24 hours

tag: genotype/variation

value: G1E cells expressing GATA1-ER


title: E18.5_WT_709_rep1

tag: strain


title: RNA_siNUDT5 rep2


title: P0_WT_704_rep2

tag: tissue


title: RNA_siNUDT5 rep1


title: E18.5_HET_710_rep1

tag: age


title: Pore_Day8 [SAM24405545]

source_name: Pore

value: Pore


title: RNA_siCtrl rep3

value: transfected with non-targeting control siRNA


title: female mESC, single clone1

description: female


title: 1663_G1E-ER4_+E2_13_hours_PRO-seq rep2

value: Estradiol (E2) for 13 hours


title: P0_HOM_701_rep1

tag: genotype

value: HOM


title: RNA_siCtrl rep2


title: P0_WT_704_rep1


title: Pore_Day8 [SAM24405543]


title: 1502_G1E-ER4_+E2_13_hours_PRO-seq rep1


title: 1866_G1E-ER4_-E2_PRO-seq_rep3


title: Mrps5 fl/fl mouse heart tissue, ctrl, embryonic 12 days, rep 2

value: Heart

value: Embryo

value: Embryonic 12 days


title: Pore_Day8 [SAM24405544]


title: 1662_G1E-ER4_-E2_PRO-seq_rep2


title: RNA_siCtrl rep1


title: Pore_Day8 [SAM24405542]


title: ATAC_siNUDT5 5Gy_rep2

data_processing:
For RNA-seq, Sequenced reads were aligned to the UCSC hg19 reference genome assembly and gene counts were quantified using STAR (v2.7.3a). Differential gene expression testing was performed by DESeq2 (v1.22.1). RNAseq analysis was performed using the VIPER snakemake pipeline.
For ATAC-seq, library quality and quantification was assessed with an Agilent Bioanalyzer at the Molecular Biology Core Facilities at Dana-Farber Cancer institute (DFCI). Barcoded sample libraries were pooled and run on on Illumina NextSeq 500 (Paired-End 35bp) at the Molecular Biology Core Facilities (MBCF) at Dana-Farber Cancer institute (DFCI).
Genome_build: hg 19
Supplementary_files_format_and_content: bigwig and peak files

extract_protocol:
For ATAC-seq, 50,000 cells were lysed with ATAC-Resuspension Buffer (RSB) containing 0.1% NP40, 0.1% Tween20, and 0.01% Digitonin. Nuclei were pelleted and incubated with 100 nM Tn5 transposase (Illumina, Cat#FC-121-1030) for 30 min at 37°C. Transposed DNA was Cleaned up with Zymo DNA Clean and Concentrator-5 Kit (cat# D4014).
For ATAC-seq, The purified DNA were amplified for 3 cycles using NEBNext 2x MasterMix and purified with AMPure XP beads (Beckman).

treatment_protocol: For ATAC-seq, U2OS cells were transfected with siRNAs, 48hr later cells were treated with or without IR at 5Gy and recovered for 30 min.

value: treated with IR at 5Gy


title: Mrps5 fl/fl mouse heart tissue, ctrl, embryonic 12 days, rep 1

value: Mrps5 fl/fl


title: P22_WT_698_rep2

value: female


title: Skin_Day8 [SAM24405546]

value: Skin


title: Pore_Day8 [SAM24405541]

value: KPP-EGFP

value: Tumor


title: 1501_G1E-ER4_-E2_PRO-seq_rep1


title: 1865_G1E-ER4_+E2_24_hours_PRO-seq rep1


title: P22_WT_698_rep1

value: cortex


title: P22_HET_695_rep2

value: HET

value: male


title: Tumor_Day8 [SAM24405540]


title: 1864_G1E_PRO-seq_rep1


title: Tumor_Day8 [SAM24405539]


title: ATAC_siNUDT5 0Gy_rep2

value: transfected with siRNA targeting NUDT5


title: ATAC_siNUDT5 0Gy_rep1


title: P0_HOM_701_rep2


title: P22_HET_695_rep1


title: P0_HET_693_rep1


title: ATAC_siCtrl_5Gy_rep2

value: untreated


title: 2538_G1E-GATA1(V205M)-ER_+E2_24_hours_H3K27ac_rep1

data_processing:
Bowtie 1.2.1.1 was used to align sequences. Reads with more than one mismatch or multiple alignments were excluded.
Significantly enriched regions were called using MACS2 version 2.1.0 with the following parameters: P = 10−5, extsize = 300 and local lambda = 100,000 using whole-cell extract input controls.
Reads were RPM normalized for  bigwigs.
Genome_build: mm9
Supplementary_files_format_and_content: bigwig files

source_name: G1E-ER4 V205M

growth_protocol: G1E, G1E-ER4 and G1E-GATA1(V205M)-ER cells were grown in IMDM media (Corning, 10-016-CV) with 15% fetal calf serum (Gemini, lot A08H00K), 2% penicillin/streptomycin (Gibco, 15140-122), 140 μm 1-thioglycerol (Sigma, M6145), 2 units/ml epoetin (Amgen) and conditioned medium from a kit ligand-producing CHO cell line. Cells were grown at a cell density of 100,000 to 1,000,000 cells/ml.

extract_protocol:
Cells were crosslinked in 1% formaldehyde in PBS/2mM EDTA at room temperature for 10 minutes. Reaction was quenched with glycine. Cell pellets were washed in PBS, resuspended in 1 ml cell lysis buffer (Methods) and incubated on ice for 20 minutes. Nuclei were pelleted and resuspended in 1 ml nuclear lysis buffer, incubated on ice for 20 minutes and sonicated for 45 minutes at 4 degrees Celsius. After sonication, samples were spun at 21130×g for 5 minutes at 4C. Supernatant (~ 1 ml) was diluted using 4 ml IP Dilution Buffer (Methods). Samples were precleared using 50 ug of isotope-matched IgG and 50 ul A/G agarose beads. IP reactions were set up by adding A/G beads pre-bound with antibody of interest and rotated overnight at 4 degrees Celsius. On the second day, beads were washed and eluted twice with 100 ul Elution Buffer. After reverse crosslinking, DNA was purified using the QIAquick PCR Purification kit per the manufacturer’s instructions.
ChIP-seq libraries were prepared using Illumina’s TruSeq ChIP sample preparation kit (Illumina, Cat#IP-202-1012) according to manufacturer’s protocol.

value: G1E-ER4 V205M

value: Estradiol (E2) for 24 hours

tag: antibody

value: Active Motif #39133

value: G1E cells expressing GATA1(V205M)-ER


title: Tumor_Day8 [SAM24405538]


title: ATAC_siCtrl_5Gy_rep1

value: transfected with non-targeting control siRNA


title: 2536_G1E-GATA1(V205M)-ER_+E2_13_hours_H3K27ac_rep2

value: Estradiol (E2) for 13 hours


title: Tumor_Day8 [SAM24405536]


title: 2537_G1E-GATA1(V205M)-ER_+E2_24_hours_H3K27ac_rep2


title: ATAC_siCtrl_0Gy_rep2


title: Tumor_Day8 [SAM24405537]


title: P0_HET_693_rep2


title: P0_HOM_690_rep2

value: HOM


title: P0_HOM_690_rep1


title: 2005_G1E_PRO-seq_rep2


title: ATAC_siNUDT5 5Gy_rep1

value: treated with IR at 5Gy


title: 2076_G1E-GATA1(V205M)-ER_+E2_13_hours_H3K27ac_rep1


title: ATAC_siCtrl_0Gy_rep1


title: P22_HET_688_rep2

value: female


title: Pore_Day3 [SAM24405535]

value: Pore


title: 1344_Normal

city: New York

first: Bic

last: MSKCC

state: NY

address: 1275 York Ave.

institute: Memorial SLoan-Kettering Cancer Center

description: 1344_Normal (AR_03_Post_ControlR)

data_processing:
The demultiplexed FASTQs were aligned to human genome reference hg19/GRCh37 using STAR (version 2.7.3a) and deduplicated from UMI and alignment coordinate using UMI-tools (version 1.0.1).
Transcript abundance was quantified using Rsubread (version 2.4.2)
Genome_build: hg19/GRCh37
Supplementary_files_format_and_content: Read count matrix file was generated using fcount from Rsubread

source_name: R obturator LN

extract_protocol:
These are formalin-fixed paraffin embedded human tissues that underwent bulk RNA sequencing.
Approximately 200-500 ng of FFPE RNA etracted from FFPE slides with a DV200 range between 3-99 or 65-100 ng of fresh frozen RNA per sample
RNA library constructed using KAPA RNA Hyper library prep kit. Customized adapters with 3bp UMI and sample-specific dual-index primers were added. Approximately 500 ng of each RNA library were pooled for hybridization capture with IDT Whole Exome Panel V1 using a customized capture protocol modified from NimbleGen SeqCap Target Enrichment system. The captured libraries were sequenced on Illumina HiSeq4000 in paired ends (2x100bp) to a target 50 million read pairs per sample.

tag: patient number

tag: involved by tumor?

value: No

tag: region characterization

value: Normal


title: Pore_Day3 [SAM24405534]

value: KPP-EGFP

value: Tumor


title: 2075_G1E-GATA1(V205M)-ER_-E2_H3K27ac_rep1


title: Pore_Day3 [SAM24405533]


title: Pore_Day3 [SAM24405532]


title: P22_HET_688_rep1

value: cortex

value: HET

tag: Sex


title: 2534_G1E-ER4_+E2_24_hours_H3K27ac _rep2

value: G1E cells expressing GATA1-ER


title: 1344_R8

description: 1344_R8 (AR_03_Post_T8)

value: Yes

value: Viable Tumor


title: P22_HET_687_rep2


title: 1344_R7

description: 1344_R7 (AR_03_Post_T7)


title: 1344_R6

description: 1344_R6 (AR_03_Post_T6)


title: P22_HET_687_rep1


title: 1344_R5

description: 1344_R5 (AR_03_Post_T5)


title: 2533_G1E-ER4_+E2_13_hours_H3K27ac_rep2

value: Active Motif #39133


title: 1994_G1E-ER4_+E2_24_hours_H3K27ac _rep1

value: Estradiol (E2) for 24 hours


title: P22_WT_686_rep2

value: WT


title: Tumor_Day3 [SAM24405530]


title: 1993_G1E-ER4_+E2_13_hours_H3K27ac_rep1

value: Estradiol (E2) for 13 hours


title: 2535_G1E-GATA1(V205M)-ER_-E2_H3K27ac_rep2

value: G1E-ER4 V205M

value: G1E cells expressing GATA1(V205M)-ER


title: 1344_R4

description: 1344_R4 (AR_03_Post_T4)


title: 2532_G1E-ER4_-E2_H3K27ac_rep2


title: Tumor_Day3 [SAM24405528]


title: 1992_G1E-ER4_-E2_H3K27ac_rep1


title: 1344_R3

description: 1344_R3 (AR_03_Post_T3)


title: Tumor_Day3 [SAM24405527]


title: P22_WT_685_rep2

value: male


title: P22_WT_685_rep1


title: 1344_R2

description: 1344_R2 (AR_03_Post_T2)


title: 2531_G1E_H3K27ac_rep2


title: 1344_R1

description: 1344_R1 (AR_03_Post_T1)


title: adult_HET_B70_rep1

value: adult


title: 1302_Normal

description: 1302_Normal (AR_02_Post_ControlR)

source_name: LLL Lung

value: No


title: Pore_Day1 [SAM24405525]

value: Pore


title: 1991_G1E_H3K27ac_rep1


title: G1E-ER4_Kit_cDNA_-E2_0%_clone2_4C_Kit_+5kb_viewpoint

description: G1E-ER4 cells transduced with and selected for MigR1-Kit cDNA-YFP retrovirus. GATA motif within enhancer was deleted using CRISPR-Cas9 mediated genome editing

data_processing:
Library strategy: 4C-seq
4C mapping was done as described here https://github.com/deWitLab/4C_mapping
PeakC was used to call 4C peaks https://github.com/deWitLab/peakC
Genome_build: mm9
Supplementary_files_format_and_content: wig

source_name: G1E-ER4 Kit cDNA

extract_protocol: 4C-seq protocol was performed as previously described: Krijger et al., 2020. 4C-seq from beginning to end: A detailed protocol for sample preparation and data analysis.

value: GATA-1 erythroid cell

tag: sample type

value: G1E-ER4 Kit cDNA

value: No Estradiol (E2)

value: expressing GATA1-ER


title: 1302_LN9

description: 1302_LN (AR_02_Post_B1)

value: LN


title: adult_HET_B72_rep2

value: cortex

value: HET

value: female


title: Pore_Day1 [SAM24405524]

value: KPP-EGFP

value: Tumor


title: P22_WT_686_rep1

value: WT


title: G1E-ER4_Kit_cDNA_-E2_10%_clone2_4C_Kit_+5kb_viewpoint

value: transduced with and selected for MigR1-Kit cDNA-YFP retrovirus. GATA motif within enhancer was deleted using CRISPR-Cas9 mediated genome editing


title: adult_HET_B72_rep1


title: Pore_Day1 [SAM24405523]


title: Tumor_Day3 [SAM24405529]


title: 1302_R7

description: 1302_R7 (AR_02_Post_T4)

value: Yes

value: Viable Tumor


title: 1302_R8

description: 1302_R8 (AR_02_Post_B5)

value: No Viable Tumor


title: adult_HET_B57_rep2

value: adult


title: Pore_Day1 [SAM24405522]

value: Pore


title: 1302_R6

description: 1302_R6 (AR_02_Post_B4)

value: No


title: adult_HET_B57_rep1


title: G1E-ER4_Kit_cDNA_-E2_10%_clone1_4C_Kit_+5kb_viewpoint

value: GATA-1 erythroid cell

value: G1E-ER4 Kit cDNA


title: Tumor_Day1 [SAM24405520]


title: 1302_R5

description: 1302_R5 (AR_02_Post_T3)


title: Pore_Day1 [SAM24405521]


title: adult_WT_B78_rep2

value: male


title: G1E-ER4_Kit_cDNA_-E2_+5kb_viewpoint

description: G1E-ER4 cells transduced with and selected for MigR1-Kit cDNA-YFP retrovirus

value: No Estradiol (E2)

value: expressing GATA1-ER


title: Tumor_Day3 [SAM24405526]


title: Tumor_Day1 [SAM24405519]


title: Pore_Day3 [SAM24405531]


title: G1E-ER4_+E2_13_hours_4C_Atp1a1_promoter_viewpoint

description: G1E cells expressing GATA1-ER

value: Estradiol (E2) for 13 hours


title: 1302_R4

description: 1302_R4 (AR_02_Post_B3)


title: G1E_ER4_-E2_4C_Atp1a1_promoter_viewpoint

description: G1E cells expressing GATA1-ER


title: Tumor_Day1 [SAM24405518]


title: G1E-ER4_Kit_cDNA_-E2_0%_clone1_4C_Kit_+5kb_viewpoint


title: 1302_R3

description: 1302_R3 (AR_02_Post_B2)


title: G1E-ER4_+E2_13_hours_4C_Kit_+5kb_viewpoint

description: G1E cells expressing GATA1-ER


title: 1302_R2

description: 1302_R2 (AR_02_Post_T2)


title: Tumor_Day1 [SAM24405517]


title: adult_WT_B76_rep1

value: cortex

value: female


title: G1E_4C_Kit_+5kb_viewpoint

value: control


title: Tumor_Day1 [SAM24405516]

value: Tumor

value: KPP-EGFP


title: 1302_R1

description: 1302_R1 (AR_02_Post_T1)

value: Yes


title: G1E-ER4_+E2_13_hours_4C_Kit_+49kb_viewpoint

description: G1E cells expressing GATA1-ER


title: adult_WT_B58_rep1

value: WT


title: scRNA-Seq of skin [M]

city: Nanjing

first: zhang

last: zhang chenyang

address: 163 Xianlin Avenue

institute: Nanjing University

data_processing:
Sample demultiplexing, barcode processing, and single cell 3’ gene counting were performed by the Cell Ranger pipelines  (v3.1.0,10X Genomics).
Genome_build: mm10  (GRCm38)
Supplementary_files_format_and_content: barcodes.tsv; features.tsv; matrix.mtx

extract_protocol:
The cynomolgus monkey project included 3 RNA organs.
RNA: 3' transcriptome sequencing, cDNA fragmentation, end filling, adding A, connection connector, Index PCR

tag: agent

value: metformin

value: post natal 7 weeks


title: G1E_4C_Kit_+49kb_viewpoint

tag: cell type


title: 1263_Normal

description: 1263_Normal (AR_01_Post_ControlR)

value: Normal


title: scRNA-Seq of skin [S]

value: Shi-Bi-Man


title: adult_WT_B78_rep1

value: adult


title: G1E-ER4_+E2_24_hours_4C_Kit_promoter_viewpoint

description: G1E cells expressing GATA1-ER

value: GATA-1 erythroid cell

value: Estradiol (E2) for 24 hours


title: P22_HET_446_rep1

value: HET

value: male


title: scRNA-Seq of skin [C]


title: 1302_Pre-tx

description: 1302_Pre-tx (AR_02_Pre_T1)

value: Viable Tumor


title: P22_HET_446_rep2

value: cortex


title: 1263_A3

description: 1263_A3 (AR_01_Post2_T1)

source_name: Adrenal

value: Adrenal


title: 1263_A4

description: 1263_A4 (AR_01_Post2_B1)


title: G1E-ER4_+E2_13_hours_4C_Kit_promoter_viewpoint

description: G1E cells expressing GATA1-ER

value: Estradiol (E2) for 13 hours

value: expressing GATA1-ER


title: G1E_4C_Kit_promoter_viewpoint

value: No Estradiol (E2)

value: control


title: P22_HET_444_rep1


title: adult_WT_B76_rep2

value: WT

value: female


title: wild-type, RNA-seq, Replicate 2

city: Freiburg

first: Rick

last: Gelhausen

state: Baden Württemberg

address: Georges-Köhler-Allee 106

department: Department of Computer Science

institute: Albert-Ludwigs-University Freiburg

description: Total RNA extracted from WT strain, DNaseI digested, rRNA depleted, fragmented and subjected to cDNA library preparation

data_processing:
Base calling with Illumina NextSeq RTA v2.11.3
FASTQ conversion with bcl2fastq v2.20.0.422
FASTQ quality and adapter trimming using Cutadapt (Martin, 2011) version 2.1 (cutadapt parameters: -q 20 --trim-n -m 10 -a AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC) (HRIBO 1.6.0)
Mapping with segemehl 0.3.4 (HRIBO 1.6.0)
rRNA filtering was done using samtools 1.9 (HRIBO 1.6.0)
Coverage files were created using HRIBO 1.6.0
bigwig files were generated with full read coverage and normalized to the size of the minimal sample size among all samples.
Assembly: ASM34606v1
Supplementary files format and content: bigwig format files containing normalized read coverage for each sample

organism: Sinorhizobium meliloti 2011

source_name: bacterial strain

growth_protocol: For Ribo-seq sample preparation, S. meliloti cells were grown to exponential phase (OD₆₀₀ₙₘ 0.5) in minimum medium

extract_protocol:
RNA was extracted from fractions or cell pellets for total RNA using hot phenol-chloroform-isoamyl alcohol (25:24:1, Roth) or hot phenol (Roth), respectively, as described previously (Sharma et al. 2007).
Total RNA was first rRNA-depleted, fragmented then subjected to cDNA library preparation. Size selcted RNA for Riboseq samples were directly subjected to cDNA library preparation. First, an oligonucleotide adapter was ligated to the 3' end of the RNA molecules. First-strand cDNA synthesis was performed using M-MLV reverse transcriptase and the 3’ adapter as primer. The first strand cDNA was purified and the 5' Illumina TruSeq sequencing adapter was ligated to the 3' end of the antisense cDNA. The resulting cDNA was PCR-amplified to about 10-20 ng/μl using a high fidelity DNA polymerase. The cDNA was purified  using the Agencourt AMPure XP kit (Beckman Coulter Genomics) and was analyzed by capillary electrophoresis.

treatment_protocol: No treatment applied during growth.

value: bacterial strain

value: no treatment


title: 1263_LN12

description: 1263_LN12 (AR_01_Post_B3)

value: No

value: LN


title: 1263_R8

description: 1263_R8 (AR_01_Post_B7)

value: No Viable Tumor


title: wild-type, RIBO-seq, Replicate 2

description: RNA purified from WT ribosome footpints, size selected and subjected to cDNA library preparation


title: P22_HET_444_rep2


title: sfGFP-GAF(N) control 1

city: Princeton

first: Michael

last: Levine

state: New Jersey

address: Icahn lab, Princeton University

institute: Princeton University

description:
embryo_loop_list.bedpe
embryo_contact_count_table.txt
embryo_differential_looping.xlsx

library_source: other

data_processing:
The raw data were mapped onto the Dm6 reference genome as described by bwa (Li and Durbin, 2009).
Validated paired reads were filtered and finalized using pairtools (Goloborodko et al., 2019)
They were further processed into multi-resolution matrix by Cooler (Abdennur and Mirny, 2020)
Focal contacts in wing imaginal discs expressing Halo-GAF were detected by SIP (Rowley et al., 2020) using newly created Micro-C maps. The parameters were set as -g 4.0 -min 2.0 -max 2.0 -mat 5000 -d 15 -res 400 -sat 0.01 -t 2800 -nbZero 4 -factor 2 -fdr 0.1 -del true -cpu 16 -isDroso false. The called loops were further filtered with a “value” > 4.25 and with enough micro-C data coverage (filtered out anchors with marginals < 500). The loops were further filtered by removing loops associated with high CP190 (>10000) (Kyrchanova et al., 2021) by intersecting with BEDtools (Quinlan and Hall, 2010). The loops were first mapped to transcription start sites within 2500 bp of the loop anchor center using BEDtools (Quinlan and Hall, 2010). They were subsequently manually inspected to determine whether linked promoters correspond to paralogs.
The dumped matrix with 100bp bin size from the cooler files were compared with the standard loop list with anchors normalized to 1.6kb for embryos and 3.2kb for discs using the pairToPair function from BEDtools (Quinlan and Hall, 2010).
The reads were summed for each standard loop and summarized into a count table. The count tables were imported for differential looping analysis with DESeq2 (Love et al., 2017).
Dm6
bedpe, embryo loop list from Levo et al., 2022
Excel, DEG result of  Micro-C contact counts
cooler, contact matrix
Library strategy: Micro-C

molecule: other

organism: Drosophila melanogaster

source_name: 2-2.5h embryo

extract_protocol:
GAF deGradFP depleted embryos and controls were collected on apple juice agar plate and harvested 2–2.5 hr after fertilization. Embryos were dechorionated for 2 min in 50% bleach and rinsed with water. Initial fixation was performed in 3.5 ml PBST (0.1% Triton-X in PBS), 6.5 ml N-heptane and 1 ml fresh 16% formaldehyde in a horizontal shaker for 15 min at 250 rpm. 3.7 ml of 2 M Tris-HCl pH 7.5 was added to terminate fixation
The fixation was terminated by addition of 370 µL 2 M Tris-HCl pH 7.5. Secondary fixations with 3 mM DSG (Thermo Fisher Scientific) and EGS (Thermo Fisher Scientific) were performed as previously described (Levo et al., 2022).

value: 2-2.5h embryo

value: sfGFP-GAF(N)


title: wild-type, RNA-seq, Replicate 1


title: sfGFP-GAF(N) control 2


title: adult_WT_B58_rep2

value: adult


title: 1263_R7

description: 1263_R7 (AR_01_Post_T4)

value: Yes

value: Viable Tumor


title: wild-type, RIBO-seq, Replicate 1


title: sfGFP-GAF(N) control 3


title: iPSC_MBE1006

type: genomic

city: Darlinghurst

first: Drew

last: Neavin

state: New South Wales

address: 384 Victoria St

department: Cellular Science

country: Australia

institute: Garvan Institute of Medical Research

data_processing:
SNPs were filtered for low-quality SNPs in Axiom Suite. Genotyped SNPs were then imputed using hg38 1000G v3 using an in-house pipeline described and available on github: https://github.com/powellgenomicslab/SNP_imputation_1000g_hg38/wiki/SNP-Genotype-Imputation-Using-1000G-hg38-Reference.
Processed data is provided in two vcf files - one that contains the iPSC lines in the 18-line village (18 of the samples) and one that contains iPSC lines from the 3-line village (3 samples)

label: biotin

source_name: iPSCs

growth_protocol: 1mL aliquots of each of the iPSC line were thawed and plated in StemFlex media (Life Technologies; Catalog Number: A3349401; Lot Number: 2093181) complete with StemFlex Supplement (Life Technologies, Catalog Number: A33492-01; Lot Number: 2090179) with Rock inhibitor Y-27632 (10µM final concentration; Stem Cell Technologies; Catalog Number: 72304) on Costar non-treated 6 well polystyrene plates (Catalog Number: 3736; Lot Number: 30417038) coated with Vitronectin XF (Stem Cell Technologies; Catalog Number: 07180; Lot Number: 18B87584) diluted in CellAdhere Buffer (Stem Cell Technologies, Catalog Number: 07183; Lot Number: 18M979058). Cells were subcultured once a week with ReLeSR (Stem Cell Technologies; Catalog Number: 05872) until sample collection.

extract_protocol: DNA was extracted with the QIAGEN DNeasy Blood & Tissue Kit

value: Male


title: 1263_R6

description: 1263_R6 (AR_01_Post_T3)


title: sfGFP-GAF(N) control 4


title: 1263_R5

description: 1263_R5 (AR_01_Post_T2)


title: P22_WT_447_rep1

value: cortex


title: 1263_R4

description: 1263_R4 (AR_01_Post_B6)


title: iPSC_WAB0103


title: 1263_R3

description: 1263_R3 (AR_01_Post_B5)


title: P22_WT_445_rep2


title: P22_HET_442_rep1

value: HET


title: 1263_R2

description: 1263_R2 (AR_01_Post_B4)


title: iPSC_WAB0004

value: Female


title: P22_WT_443_rep2

value: male


title: iPSC_WAB0038


title: iPSC_TOB0435


title: P22_WT_443_rep1

value: WT


title: 1263_R1

description: 1263_R1 (AR_01_Post_T1)


title: P22_HET_442_rep2

value: female


title: iPSC_TOB0421


title: iPSC_FSA0006


title: P22_WT_447_rep2


title: iPSC_TOB0220


title: E18.5_HOM_454_rep2

value: HOM


title: P22_WT_445_rep1


title: E18.5_HOM_454_rep1


title: control

city: Guangzhou

first: chen

middle: wei

last: fen

address: 4F, Building 1, Standard Property Unit 3, International Biotech Island

institute: Forevergen

description: circRNA-seq

data_processing:
Illumina CASAVA 1.8 software used for basecalling.
Low quality which have more than 20% low base quality bases or adapter sequence contamination reads were discarded. Then high quality reads were aligned to the reference genome using BWA mem alignment module. CircRNA identify and characterize was used the CIRI2 software based on the following process. First, reads aligning contiguously to the genome without any trimming were discarded. The remaining reads were used for circRNA candidate detection. Next, terminal sequences (anchors) were extracted and re-aligned independently to find unique anchor positions. Anchors that aligned in the reverse orientation (head-to-tail) indicated circRNA splicing (back-splicing).
The following criteria were used to filter the resulting splicing events: 1) GT/AG signal flanking the splice sites; 2) unambiguous breakpoint detection; 3) no more than two mismatches in the extension procedure; 4) the breakpoint reside a maximum of two nucleotides inside an anchor; 5) less than two independent reads support the head-to-tail splice junction.
Counts of reads mapping across an identified back-splice are normalized by number of reads mapping (spliced tags reads per million mapping, TPM), which allows expression comparisons between two circRNAs. Differential expression analysis of two samples was performed using the previous statistics model. The resulting P-values were adjusted using the Benjamini and Hochberg’s approach for controlling the false discovery rate. CircRNA with an adjusted P-value <0.05 were assigned as differentially expressed.
Assembly: rn6

organism: Rattus norvegicus

source_name: hydrostatic cancer cells

growth_protocol: Walker256 cells in the logarithmic phase were collected and washed with PBS 3 times. The cell concentration was adjusted to 1×107cells/ml and 0.5ml was injected into the peritoneal cavity of SD rats. 9-12 days later, the ascites was centrifuged to collect the Walker 256 cells and 5μl (4×105 cells) of cell suspension was injected into the bone marrow cavity 0.5cm below the tibia, while the control operation group was injected with Walker 256 cells boiled for 20 minutes and underwent the same surgical procedure as above. The bone cancer pain group and the control control group were each composed of 3 animals, which were kept for 21 days after surgery and subsequently anaesthetized and sacrificed.

extract_protocol:
Total RNA from the cells was isolated using the Trizol (Life Technologies, Carlsbad, CA, USA) according to the manufacturer’s instructions, followed by a DNase I digestion for 30 minutes to remove genomic DNA
Total RNA was treated with the RiboMinus kit (Life Technologies) to remove the ribosomal RNAs; RNAs were incubation with 16 units RNase R (Sigma–Aldrich, St. Louis, MO, USA) for 15 min at 37℃ to deplete linear and enrich circRNAs; The remaining RNAs were fragmented and reverse-transcribed using random primers and reverse transcriptase; Second strand cDNA synthesis was completed using DNA polymerase I and RNase H. Sixth, the Illumina series KAPA Library Preparation Kit (Kapa Biosystems) was used to perform end repair, dA-tailing, and adapter ligation; The dUTP-marked second strand was digested at 37 °C for 30 min using Uracil Specific Excision Reagent (USER) enzyme (New England Biolabs); The cDNA was enriched by PCR amplification to create the final cDNA libraries.

value: hydrostatic cancer cells

value: spinal cord cells from the injected rats


title: bone_cancer_pain

description: circRNA-seq


title: GAF(deGradFP) 2

value: GAF(deGradFP)


title: delta-POZ 1

description:
wing_disc_loop_list.bedpe
wing_disc_contact_count_table.txt
wing_disc_differential_looping.xlsx

data_processing:
The raw data were mapped onto the Dm6 reference genome as described by bwa (Li and Durbin, 2009).
Validated paired reads were filtered and finalized using pairtools (Goloborodko et al., 2019)
They were further processed into multi-resolution matrix by Cooler (Abdennur and Mirny, 2020)
Focal contacts in wing imaginal discs expressing Halo-GAF were detected by SIP (Rowley et al., 2020) using newly created Micro-C maps. The parameters were set as -g 4.0 -min 2.0 -max 2.0 -mat 5000 -d 15 -res 400 -sat 0.01 -t 2800 -nbZero 4 -factor 2 -fdr 0.1 -del true -cpu 16 -isDroso false. The called loops were further filtered with a “value” > 4.25 and with enough micro-C data coverage (filtered out anchors with marginals < 500). The loops were further filtered by removing loops associated with high CP190 (>10000) (Kyrchanova et al., 2021) by intersecting with BEDtools (Quinlan and Hall, 2010). The loops were first mapped to transcription start sites within 2500 bp of the loop anchor center using BEDtools (Quinlan and Hall, 2010). They were subsequently manually inspected to determine whether linked promoters correspond to paralogs.
The dumped matrix with 100bp bin size from the cooler files were compared with the standard loop list with anchors normalized to 1.6kb for embryos and 3.2kb for discs using the pairToPair function from BEDtools (Quinlan and Hall, 2010).
The reads were summed for each standard loop and summarized into a count table. The count tables were imported for differential looping analysis with DESeq2 (Love et al., 2017).
Dm6
bedpe, filtered wing disc loop list from Halo-Trl loop calling
Tab delimited table, read counts table of Micro-C contacts
Excel, DEG result of  Micro-C contact counts
cooler, contact matrix
Library strategy: Micro-C

source_name: wing imaginal disc

extract_protocol:
For wing imaginal discs, homozygous wandering 3rd instar larvae carrying Halo tagged GAF were collected and dissected in PBS. 90 discs were collected for each biological replicate and two replicates for each genotype were prepared. Discs were incubated in 500 µL 1% formaldehyde/PBST for 15 min, passively rotating at room temperature.
The fixation was terminated by addition of 370 µL 2 M Tris-HCl pH 7.5. Secondary fixations with 3 mM DSG (Thermo Fisher Scientific) and EGS (Thermo Fisher Scientific) were performed as previously described (Levo et al., 2022).

value: wing imaginal disc

value: delta-POZ


title: iPSC_TOB0205


title: Halo-Trl 1

value: halo-Trl


title: Halo-Trl 2


title: GAF(deGradFP) 3

value: 2-2.5h embryo


title: GAF(deGradFP) 4


title: GAF(deGradFP) 1


title: E18.5_HOM_461_rep2

data_processing:
Sequenced reads were trimmed for adaptor sequence and masked for low-complexity or low-quality sequence
Filtered reads were mapped to mm10 with STAR to generate raw counts files
Differential expression and normalised counts were determined using DESeq2 (R package version 1.28.1) with sequencing batch ID and sex as covariates
Genome_build: mm10
Supplementary_files_format_and_content: Tab-delimited text file with raw counts for all samples
Supplementary_files_format_and_content: Tab-delimited text file with DESeq2 output for all conditions (sample-collapsed)
Supplementary_files_format_and_content: Tab-delimited text file with normalised counts for all samples


title: iPSC_TOB0199


title: delta-POZ 2


title: E18.5_HOM_461_rep1


title: delta-Q 1

value: delta-Q


title: iPSC_TOB0198


title: E18.5_HOM_455_rep1


title: delta-Q 2


title: iPSC_MBE2900


title: E18.5_HOM_455_rep2


title: CF d120 Sheep Trachea rep 1

city: Cleveland

first: Ann

last: Harris

state: Ohio

address: 2109 Adelbert Rd Building

department: Genetics and Genome Sciences

institute: Case Western Reserve University

description:
RNA seq of d120 CF sheep trachea rep1
RNA-seq

data_processing:
Bulk RNA-seq mapped to the sheep (texel, Oar_v4.0) genome with STAR 2.6 (https://github.com/alexdobin/STAR)
Aligned reads were assigned to genomic features with featureCounts version 1.6.3 in the Subread package (http://subread.sourceforge.net/) to estimate expression levels for each transcript.
Assembly: Texel (ovis aries) sheep genome Oar_v4.0
Supplementary files format and content: Feature assignment of aligned reads to annotated sheep genes in TSV format.

organism: Ovis aries

source_name: sheep tracheal epithelial tissue

extract_protocol:
50-100mg of sample was excised from frozen tissue and mixed with 1mL TRIzol Reagent and ceramic beads over ice. Tissue was homogenized and incubated at RT to dissociate the nucleoprotein complex. Solution was then mixed with 200uL chloroform, centrifuged, and the aqueous phase removed. RNA was precipitated with 100% isopropanol, washed with 75% ethanol, and solubilized in water.
RNA concentration and purity were measured by nanodrop and Tapestation and 1ug was used for library preparation.

tag: subject

value: CF donor

value: Day 120

tag: sample


title: Halo-Trl CnT 1

data_processing:
CUT&Tag data was processed using an integrated analysis pipeline for CUT&RUN and CUT&Tag, nf-core/cutandrun v2.0 (Cheshire and West), with relax peak calling using SEACR (Meers et al., 2019).
Dm6
bigWig, normalized counts of CUT&Tag result
Library strategy: CUT&Tag

extract_protocol:
12 wing discs from homozygous wandering 3rd instar larvae of Halo-GAF lines were collected for each biological replicate and two replicates for each genotype were prepared.
The discs were incubated with 1:100 rabbit polyclonal Halotag antibody (Promega G9281) for Halo-GAF lines, and 12 wildtype discs were incubated with 1:100 rabbit anti-mouse IgG H&L (Abcam ab46540) as an IgG control. Ginea pig α-rabbit antibody (Antibodies online ABIN101961) was used at 1:100 dilution as secondary antibody. 0.1ng of E. coli Spike-in DNA (EpiCypher 18-1401) was added to each sample at the same step where pAG-Tn5 (EpiCypher 15-1117) was added. DNA was purified by phenol-chloroform extraction and ethanol precipitation, then universal i5 primers and uniquely barcoded i7 primers were added to the DNA with 14 cycles of PCR. Individual libraries were purified with Ampure XP beads (Beckman Coulter A63880) at 1.3:1 ratio and eluted with 22 μL 10mM Tris-HCL, pH8. Mixed barcoded libraries were purified again with Ampure XP beads at 1.8:1 ratio and eluted with 20 μL μL 10mM Tris-HCL, pH8, the libraries were concentrated and the residual primers were eliminated after this step.


title: iPSC_MBE2817

value: Male


title: CF d147 Sheep Trachea rep 2

description:
RNA seq of d147 (birth) CF sheep trachea rep2
RNA-seq

value: Day 147 (birth)


title: CF d100 Sheep Trachea rep 2

description:
RNA seq of d100 CF sheep trachea rep2
RNA-seq

value: Day 100


title: E18.5_HET_457_rep2

extract_protocol:
The cortex of E18.5 or the prefrontal cortex of P0, P22, and adult (3-month-old) mice was dissected and placed into TRIzol (Invitrogen) followed by RNA harvest using RNA Miniprep kit (Zymo Research).
RNA libraries were prepared for sequencing using standard Illumina protocols

value: cortex

value: HET


title: Halo-Trl CnT 2


title: iPSC_MBE0953


title: iPSC_IST3323


title: CF d100 Sheep Trachea rep 1

description:
RNA seq of d100 CF sheep trachea rep1
RNA-seq


title: iPSC_IST1877

value: Female


title: E18.5_HET_457_rep1

value: female


title: CF d65 Sheep Trachea rep 2

description:
RNA seq of d65 CF sheep trachea rep2
RNA-seq

value: Day 65


title: E18.5_HET_460_rep2

value: male


title: delta-POZ CnT 2


title: E18.5_HET_459_rep1


title: CF d120 Sheep Trachea rep 2

description:
RNA seq of d120 CF sheep trachea rep2
RNA-seq


title: CF d80 Sheep Trachea rep 2

description:
RNA seq of d80 CF sheep trachea rep2
RNA-seq

value: Day 80


title: iPSC_FSA0004


title: delta-Q CnT 1

value: wing imaginal disc


title: CF d80 Sheep Trachea rep 1

description:
RNA seq of d80 CF sheep trachea rep1
RNA-seq


title: delta-POZ CnT 1

value: delta-POZ


title: delta-Q CnT 2


title: iPSC_FSA0001


title: CF d65 Sheep Trachea rep 1

description:
RNA seq of d65 CF sheep trachea rep1
RNA-seq


title: E18.5_HET_459_rep2


title: E18.5_HET_460_rep1


title: iPSC_180N


title: CnT input

value: halo-Trl


title: E18.5_WT_456_rep2

value: WT


title: adult_HET_B70_rep2

value: adult


title: WT d147 Sheep Trachea rep 2

description:
RNA seq of d147 (birth) WT sheep trachea rep2
RNA-seq

value: healthy donor


title: Halo-Trl RNA-seq 1

description:
RNA-seq_count_table.txt
RNA-seq_DEG.xlsx

data_processing:
The sequence was aligned to Dm6 with RNASTAR (Widmann et al., 2012)
count table was generated by featureCounts (Liao et al., 2014).
The differential gene analysis was performed with DESeq2 (Love et al., 2017)
Dm6
Tab delimited table, read counts of RNA-seq
Excel, DEG result of RNA-seq

extract_protocol:
30 wing imaginal discs were dissected from wandering 3rd instar larvae of Halo tagged lines in PBS and transferred to TRIzol (Thermo Fisher Scientific) for each biological triplicate. The total RNA was extracted based on standard protocols.
The library preparation and sequencing were performed by the Princeton University Genomics Core Facility.


title: WT d147 Sheep Trachea rep 1

description:
RNA seq of d147 (birth) WT sheep trachea rep1
RNA-seq


title: Halo-Trl RNA-seq 2


title: iPSC_166


title: WT d120 Sheep Trachea rep 2

description:
RNA seq of d120 WT sheep trachea rep2
RNA-seq

value: Day 120


title: WT d120 Sheep Trachea rep 1

description:
RNA seq of d120 WT sheep trachea rep1
RNA-seq


title: E18.5_WT_449_rep1


title: delta-POZ RNA-seq 1


title: Halo-Trl RNA-seq 3


title: WT d100 Sheep Trachea rep 2

description:
RNA seq of d100 WT sheep trachea rep2
RNA-seq


title: P0_HOM_324_rep1

value: HOM


title: P0_HOM_318_rep1


title: WT d100 Sheep Trachea rep 1

description:
RNA seq of d100 WT sheep trachea rep1
RNA-seq


title: delta-POZ RNA-seq 2


title: delta-POZ RNA-seq 3


title: E18.5_WT_449_rep2


title: Compound 8 (8-73) 10 µM-treated Mycobacterium marinum replicate 1

city: Amsterdam

first: Vien

last: Ho

state: Noord-Holland

address: De Boelelaan 1108, O2 building

department: Medical Microbiology and Infection Control

country: Netherlands

institute: Amsterdam University Medical Center-location VU

data_processing:
CLC Genomics Workbench v 12 or bcl2fastq2 version 2.18/Tophat2 version 2.1.1
Sequence reads were trimmed for adaptor sequence/low-quality sequence using CLC genomic benchwork or  bcl2fastq2 version 2.18
Trimmed sequence reads were mapped to GrCM38/mm10 using CLC genomic benchwork or Tophat2 version 2.1.1 (
Read count extraction and normalization were performed using CLC genomic benchwork
Assembly: ASM1834v1
Supplementary files format and content: tab-delimited text files include fpkm values for each Sample

organism: Mycobacterium marinum

source_name: M strain

growth_protocol: Mycobacterium marinum was grown in 7H9 medium supplemented with ADS (supplement ADS (BSA 50 g/L, dextrose 20 g/L, NaCl 8,1 g/L) and 0.02% tyloxapol

extract_protocol:
The RNA samples were isolated using the NucleoSpin RNA kit (Macherey-Nagel). The bacteria were lysed by bead beating with 0.1-zirconium beads in 500 μL buffer RA1 and 5 μL β-mercaptoethanol for 1 min. All subsequent steps followed the recommendations of the manufacturer. The rRNA was depleted and mRNA was enriched using RiboMinus prokaryotic kit (Invitrogen).
Conducted by BaseClear (Leiden, Netherlands)

treatment_protocol: Compound 8-73 or DMSO was added to culture at 10 µM or 20 µM with the bacteria starting from OD 0.05

value: M strain

tag: treatment

value: Compound 8 (8-73)


title: WT d80 Sheep Trachea rep 2

description:
RNA seq of d80 WT sheep trachea rep2
RNA-seq


title: DMSO 20 µM-treated Mycobacterium marinum replicate 2

value: DMSO


title: Compound 8 (8-73) 20 µM-treated Mycobacterium marinum replicate 1


title: delta-Q RNA-seq 1

value: delta-Q


title: Compound 8 (8-73) 10 µM-treated Mycobacterium marinum replicate 2


title: Compound 8 (8-73) 20 µM-treated Mycobacterium marinum replicate 2


title: E18.5_WT_456_rep1

source_name: mouse cortex

value: cortex


title: DMSO 20 µM-treated Mycobacterium marinum replicate 1


title: DMSO 10 µM-treated Mycobacterium marinum replicate 2


title: delta-Q RNA-seq 2


title: WT d65 Sheep Trachea rep 2

description:
RNA seq of d65 WT sheep trachea rep2
RNA-seq


title: WT d80 Sheep Trachea rep 1

description:
RNA seq of d80 WT sheep trachea rep1
RNA-seq


title: P0_WT_311_rep1


title: delta-Q RNA-seq 3


title: P0_HET_317_rep1

value: HET


title: CF d147 Sheep Trachea rep 1

description:
RNA seq of d147 (birth) CF sheep trachea rep1
RNA-seq

value: CF donor

value: Day 147 (birth)


title: WT d65 Sheep Trachea rep 1

description:
RNA seq of d65 WT sheep trachea rep1
RNA-seq

value: Day 65


title: P0_WT_309_rep1

value: male


title: P0_HET_306_rep1

value: female


title: DMSO 10 µM-treated Mycobacterium marinum replicate 1


title: 4-5 hr replicate 1

status: Public on Feb 13 2023

city: Aurora

first: Kent

middle: Augustus

last: Riemondy

state: Colorado

department: Biochemistry and Molecular Genetics

institute: University of Colorado School of Medicine

data_processing:
Library strategy: Direct RNA-seq
Base calling was performed using Guppy (v. 3.1.5+781ed57) using default settings. Sequencing reads from n = 2 replicate libraries from each time point were aligned to the Drosophila transcriptome and independently to the genome using minimap2. Transcript abundance was quantified using Salmon. Reads counts from each time point were combined.  Genome alignments were used for visualizing read coverage with each read assigned to a transcript based on the transcriptome alignment.
Genome_build: Ensembl BDGP6 release 84
Supplementary_files_format_and_content: Salmon quantification output files

source_name: embryo

growth_protocol: Embryos were collected at various time points post-egg laying, dechorionated with bleach, and washed with 0.1% Triton X-100.

extract_protocol:
Embryos were then homogenized in lysis Buffer B (150 mM KCl, 20 mM HEPES-KOH pH 7.4, 1 mM MgCl2, 1mM DTT, complete mini EDTA-free protease inhibitors), and were clarified at 15,000 rpm, 4°C for 15 minutes. The supernatant was stored at –80°C. RNA was extracted using TRI-reagent, according to the manufacturer’s instructions.
Nanopore libraries were prepared according to the Direct RNA sequencing protocol (SQK-RNA002). Because the lengths of poly(A) tails change during the MZT, total RNA was used in place of poly(A) selected RNA. Libraries were sequenced on a FLO-MIN106 flow cell and minION sequencing device.

tag: developmental stage

value: 4-5 hr AEL

value: embryo


title: 0-1 hr replicate 1

value: 0-1 hr AEL


title: 4-5 hr replicate 2


title: 0-1 hr replicate 2


title: Benign Breast Diseases [BC1448]

type: RNA

city: Kamakura

first: Ichikawa

last: Makiko

state: Kanagawa

country: Japan

institute: Toray Industries, Inc.

description: miRNA

hyb_protocol: Hybridized for 16 h at 32 ºC with rotary shaker (250 rpm). Hybridization buffer and washing protocol were attached in the 3D-Gene® miRNA oligo chip kit.

scan_protocol: 3D-Gene® Scanner (Toray Industries, Inc.) was used for scanning. Images were quantified using Extraction version 2.0.0.6 (Toray Industries, Inc.).

data_processing: The presence of miRNA was determined based on a corresponding microarray signal of greater than [the mean + 2 × standard deviation] of the negative controls signal, of which the top and bottom ranked ones by signal intensity were removed. Once a miRNA was considered present, the mean signal of the negative controls of which the top and bottom 5% ranked by signal intensity were removed was subtracted from the miRNA signal. When the signal value was negative (or undetEsophageal Cancerted) after background subtraction, the value was replaced by 0.1 on a base 2 logarithm scale.

source_name: Serum

label_protocol: miRNA was labeled using 3D-Gene® miRNA Labeling kit in accordance with the manufacturer's instructions.

extract_protocol: Total RNA was extracted each from 300uL serum samples using 3D-Gene® RNA extraction reagent from liquid sample kit (Toray Industries, Inc.).

tag: disease status

value: Benign Breast Disease

value: Serum

tag: gender

value: Female

tag: tumor stage (the pathological diagnosis of breast cancer

value: TNM Classification 8th edition):NA

tag: lymphovascular invasion (the pathological diagnosis of breast cancer)

tag: suspicious lymph node metastasis (ultrasound finding)


title: Benign Breast Diseases [BC1347]

description: miRNA

tag: tissue

tag: age


title: Benign Breast Diseases [BC1621]

description: miRNA


title: Benign Breast Diseases [BC0950]

description: miRNA


title: Benign Breast Diseases [BC1453]

description: miRNA


title: Benign Breast Diseases [BC0846]

description: miRNA


title: Benign Breast Diseases [BC1451]

description: miRNA


title: Benign Breast Diseases [BC1284]

description: miRNA


title: Benign Breast Diseases [BC1299]

description: miRNA


title: Benign Breast Diseases [BC0845]

description: miRNA


title: Benign Breast Diseases [BC1348]

description: miRNA


title: Benign Breast Diseases [BC1154]

description: miRNA


title: Benign Breast Diseases [BC1445]

description: miRNA


title: Benign Breast Diseases [BC1164]

description: miRNA


title: Benign Breast Diseases [BC1433]

description: miRNA


title: Benign Breast Diseases [BC1366]

description: miRNA


title: Benign Breast Diseases [SA0806]

description: miRNA


title: Benign Breast Diseases [BC1509]

description: miRNA


title: Benign Breast Diseases [BC0952]

description: miRNA


title: Benign Breast Diseases [BC1305]

description: miRNA


title: Benign Breast Diseases [BC1587]

description: miRNA


title: Benign Breast Diseases [BC1461]

description: miRNA


title: Benign Breast Diseases [BC1094]

description: miRNA


title: Benign Breast Diseases [BC1110]

description: miRNA


title: Benign Breast Diseases [BC1123]

description: miRNA


title: Benign Breast Diseases [BC1111]

description: miRNA


title: Benign Breast Diseases [BC1269]

description: miRNA


title: Benign Breast Diseases [BC0801]

description: miRNA


title: Benign Breast Diseases [BC1405]

description: miRNA


title: Benign Breast Diseases [BC1253]

description: miRNA


title: Benign Breast Diseases [BC1089]

description: miRNA


title: Benign Breast Diseases [BC0977]

description: miRNA


title: Benign Breast Diseases [BC0962]

description: miRNA


title: Benign Breast Diseases [BC0973]

description: miRNA


title: Benign Breast Diseases [BC0983]

description: miRNA


title: Breast Cancer [BC1277]

description: miRNA

value: Breast Cancer

value: TNM Classification 8th edition):T1

value: No


title: Breast Cancer [BC0590]

description: miRNA

value: TNM Classification 8th edition):T2


title: Breast Cancer [BC1578]

description: miRNA


title: Breast Cancer [BC0577]

description: miRNA

value: Yes


title: Breast Cancer [BC0238]

description: miRNA


title: Breast Cancer [BC0865]

description: miRNA


title: Breast Cancer [BC1147]

description: miRNA


title: Breast Cancer [BC1068]

description: miRNA


title: Breast Cancer [BC0068]

description: miRNA


title: Breast Cancer [BC0321]

description: miRNA


title: Breast Cancer [BC0834]

description: miRNA


title: Breast Cancer [BC0202]

description: miRNA


title: Breast Cancer [BC0502]

description: miRNA


title: Breast Cancer [BC0195]

description: miRNA


title: Breast Cancer [BC0182]

description: miRNA


title: Breast Cancer [BC1246]

description: miRNA


title: Breast Cancer [BC1303]

description: miRNA

value: TNM Classification 8th edition):Tis


title: Breast Cancer [BC0020]

description: miRNA


title: Breast Cancer [BC1232]

description: miRNA


title: Benign Breast Diseases [BC0921]

description: miRNA


title: Breast Cancer [BC1354]

description: miRNA


title: Breast Cancer [BC0409]

description: miRNA


title: Breast Cancer [BC0619]

description: miRNA


title: Benign Breast Diseases [BC1053]

description: miRNA


title: Breast Cancer [BC0653]

description: miRNA


title: Breast Cancer [BC0065]

description: miRNA


title: Breast Cancer [BC1122]

description: miRNA


title: Breast Cancer [BC0808]

description: miRNA


title: Breast Cancer [BC0944]

description: miRNA


title: Breast Cancer [BC0923]

description: miRNA