title: Normal5_Expr_raw

city: Los angeles

first: Xue

last: Liu

state: CA

address: AHSP A9404, 127 S. San Vicente Blvd

department: Medicine

institute: Cedars-Sinai Medical Center

data_processing:
We used Cell Ranger version 1.3.1 (10x Genomics) to process raw sequencing data and Cell Ranger R kit version 3.0.0 and Seurat suite version 3.0.0 for downstream analysis.
For clustering, principal-component analysis was performed for dimension reduction.
Top 10 principal components (PCs) were selected by using a permutationbased test implemented in Seurat and passed to t-SNE for clustering visualization. sSeq version 1.0.0 integrated in the Cell Ranger R kit was used for modeling the gene expression with negative binomial distribution to identify genes whose expression was enriched in specific clusters. The Benjamini-Hochberg procedure was used for correcting errors of multiple testing.

source_name: Healthy donor lung

extract_protocol:
Freshly isolated human lung tissues were obtained from Cedars-Sinai Medical Center and UCLA and were dissociated using a standard protocol in our laboratory64. Lung tissue was taken to a sterile tissue culture hood, transferred to a Petri dish and rinsed in PBS. Airways >2 mm were resected from the surrounding tissues and discarded along with the visceral pleura. The remaining tissue was finely minced with a scissors and then a straight razor blade. The minced lung tissue was then washed in Ham’s/F12 media (#11320033, Gibco, Thermo Fisher Scientific, Waltham, MO, USA) at 4C for 20 minutes to remove blood and then centrifuged at 600 rpm for 5 minutes in a pre-cooled centrifuge. The media was removed, and the tissue transferred to a 50 ml conical tube containing 2 mg/ml Dispase II (Thermo Fisher Scientific, Waltham, MA, USA) in DMEM/F12 overnight at 4C with gentle agitation. The next day the suspension was heated to 37C for 30 minutes, and the centrifuged for 5 minutes at 4C. The supernatant was removed, and any large pieces of tissue finely minced again with a straight razor blade. The tissue was then titurated in a digestion media containing 10 U/ml elastase (#LS002280, Worthington Biochemical, Lakewood, NJ, USA) and incubated for 30 minutes at 37C. An equal volume of HBSS+ was then added, the solution titurated and then centrifuged at (600 g, 5 minutes, 4C). The supernatant was removed, and the tissue incubated at 37C for 15 minutes with DNase I solution. The suspension was titurated and transferred to a 70 m cell strainer over a new 50 ml tube. The strainer was rinsed three times with 10 ml HBSS+. The suspension was centrifuged (600 g, 5 minutes, 4C) and the cells resuspended in 1 X RBS lysis buffer for 2 minutes on ice, the solution diluted with HBSS then centrifuged (600 g, 5 minutes, 4C). The supernatant was removed, and the cells resuspended in appropriate solution for further analysis.
mRNA from single cells sorted from lung into lysis plates was reverse transcribed to complementary DNA (cDNA) and amplified as previously described. Library preparation and sequencing were performed as described previously. Sequencing libraries for cDNA from single cells were prepared as per the Single Cell 3′ v2 Reagent Kits User Guide (10x Genomics, Pleasanton, CA, USA). Cellular suspensions were loaded on a Chromium Controller instrument (10x Genomics) to generate single-cell Gel Bead-In-EMulsions (GEMs). GEM-reverse transcription (RT) was performed in a Veriti 96-well thermal cycler (Thermo Fisher Scientific, Waltham, MA, USA). GEMs were collected and the cDNA was amplified and purified with SPRIselect Reagent Kit (Beckman Coulter, Brea, CA, USA). Indexed sequencing libraries were constructed using Chromium Single-Cell 3′ Library Kit for enzymatic fragmentation, end-repair, A-tailing, adapter ligation, ligation cleanup, sample index PCR, and PCR cleanup. The barcoded sequencing libraries were quantified by quantitative PCR using the KAPA Library Quantification Kit for Illumina platforms (KAPA Biosystems, Roche Holding AG, Basel, Switzerland). Sequencing libraries were loaded on a NextSeq500 (Illumina, San Diego, CA, USA) with a custom sequencing setting (26bp for Read 1 and 98bp for Read 2) to obtain a sequencing depth of ~200K reads per cell.

tag: mouse lung cells

value: Sorted human lung mesenchymal cells

value: Healthy

tag: assay

value: scRNA-sequencing


title: PSK3: Planktonic-free_Rep3

status: Public on Jun 30 2021

type: SRA

city: Khon Kaen

first: Supaksorn

last: Chattagul

state: Khon Kaen

address: 123 Moo 16 Mittraphap Rd., Nai-Muang

department: Biochemistry

country: Thailand

institute: Khon Kaen University

library_source: transcriptomic

data_processing:
The RNA-Seq libraries were sequenced using a single-end reads mode with an Illumina HiSeq 1500 platform (Illumina, USA). The raw RNA-Seq data were formatted in the FASTQ file. The FASTQ datasets were subjected to evaluate the quality control with FastQC [v0.11.4], and then the reads were trimmed by using Trimmomatic [v0.35] with a leading and trailing minimum score of 3 and a 4-base sliding window minimum score of 15. Post-trimmed reads were aligned to a reference of the chromosome I and II of the B. pseudomallei strain K96243 (GenBank accession no. NC006350.1 and NC006351.1) (NCBI, NIH) using Bowtie2 [v2.1.0]. The differential of gene-levels count was determined with Cuffdiff, a part of the Cufﬂinks package [v2.1.1], and a total of the normalized-genes output was reported as fragments per kilobase per million mapped reads (FPKM) with q-value of < 0.05 and 2-fold change above 1 (Log2 scale).
Genome_build: NC006350.1 and NC006351.1
Supplementary_files_format_and_content: Cuffdiff files include FPKM values for each Sample ...
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

molecule: total RNA

organism: Burkholderia pseudomallei K96243

source_name: Planktonic-free cell

growth_protocol: Overnight bacterial culture was grown in the enrichment liquid Luria-Bertani (LB) broth at 37°C with shaking until the exponential phase was reached. Planktonic-free cell pellets were recovered by centrifugation at 8,000×g for 5 min. Bacterial biofilm cells were cultured in the modified Vogel and Bonner’s medium (MVBM), in 24-well plates (Thermo Fisher Scientific, USA). An initial inoculum was diluted in MVBM to obtain 1.0×107 CFU mL-1 and grown at 37°C with shaking at 100 rpm for 24 hr. Bacterial biofilm was rinsed with phosphate buffered saline (PBS, pH 7.4) for 3 times and bacterial cells were collected by scrapping in TRIzol reagent. Planktonic shedding cells were obtained from MVBM culture using the modification of Nunc-Immuno TSP with 96 pins at the lid (Thermo Fisher Scientific, USA), Bacterial biofilm that formed was rinsed with PBS for 3 times and transferred the lids to 96-wells plate that contained enrich medium and incubated at 37°C for another 24 hr. The cell pellets in the medium were processed the same as planktonic cell and cultured in LB for 24 hr.

extract_protocol:
Total RNA was extracted from bacterial pellets and performed using a TRIzol reagent with the Purelink RNA mini kit (Ambion, USA). RNA integrity was evaluated using Agilent 2100 Bioanalyzer (Agilent Technologies, USA). The total RNA sample 2.5 μg was used to construct the RNA libraries. Total RNA in each sample, after removal of the 23S and 16S ribosomal RNA (rRNAs) with Ribo-Zero Gram-negative Bacteria rRNA removal kit (Epicentre Technologies, USA), was purified by an ethanol precipitation. The cDNA libraries were assembled from the fragmented purified rRNA-depleted RNA using a ScriptSeq v2 RNA-Seq Library Preparation kit (Epicentre Technologies, USA) and purified using a QIAquick PCR purification kit (QIAGEN, USA). An Illumina Indexed-adapter sequence was ligated to di-tagged cDNA to generate the amplified libraries as per the RNA sample preparation protocol (Illumina, USA). Excess nucleotides and PCR primers were eliminated from the libraries with a QIAquick PCR purification kit and the quantity and quality were examined using an Agilent 2100 Bioanalyzer.
cDNA libraries were prepared and constructed using a ScriptSeq v2 RNA-Seq Library Preparation kit (Epicentre Technologies, USA)

tag: strain

tag: growth condition

value: Mid-log planktonic-free cell growing in Luria-Bertani broth (LB)


title: PSK1: Planktonic-free_Rep1


title: B16-F10.ms023_1uM.I

city: Seattle

first: James

middle: Douglas

last: Thomas

state: WA

address: 1100 Fairview Ave. N., B3-111

department: Computational Biology

country: USA

institute: Fred Hutchinson Cancer Research Center

description: RNA-seq of cell line treated with the indicated compound

data_processing:
Illumina Casava software was used for base calling.
For RNA-seq, reads were aligned to the mouse transcriptome using RSEM followed by mapping to the mouse genome (mm10) using Bowtie and TopHat.
RNA splicing was quantified using MISO.
Genome_build: mm10 for mouse data
Supplementary_files_format_and_content: The final processed data is found in the file "d.psi.txt"

molecule: polyA RNA

organism: Mus musculus

extract_protocol:
RNA was extracted from cells of interest using an RNeasy kit (Qiagen, Frederick, MD) and quantified using a NanoDrop 8000 (ThermoFisher Scientific); a minimum of 500 ng of high quality RNA (as determined by Agilent Bioanalyzer) per sample or replicate was used for library preparation.
Poly(A)-selected, stranded Illumina libraries were prepared with a modified TruSeq protocol and sequenced on the Illumina HiSeq 2000

treatment_protocol: Cell lines were treated with splicing inhibitors at the indicated concentrations for 96 hours in vitro, harvested and washed three times with PBS in excess to remove all drug, and then used for downstream analyses, continued growth in vitro or tumor challenge in vivo into syngeneic animals.

tag: cell line


title: G011

city: Utrecht

first: Michael

last: VanInsberghe

state: Utrecht

address: Uppsalalaan 8

country: Netherlands

institute: Hubrecht Institute

description:
processed_data_file: RPE-FUCCI_metadata.csv.gz
processed_data_file: RPE-FUCCI_RPFcounts.csv.gz
processed_data_file: RPE_G0_layout.csv.gz

data_processing:
See github repository https://github.com/mvanins/scRiboSeq_manuscript for code and detailed instructions.
Basecalls were performed on the Illumina NextSeq 500. Reads from each plate were demultiplexed using bcl2fastq (version 2.20.0.422) with --use-bases-mask Y*,I*,Y* --no-lane-splitting --mask-short-adapter-reads 0 --minimum-trimmed-read-length 0. Read 1 contains the insert and Read 2 contains the cell barcode.
Next, the UMI was extracted from the first 10 bases of read 1 and concatenated to the start of the cell barcode. Adapter sequences were then trimmed from read 1 using cutadapt (version 2.7) with -m 15: -a TGGAATTCTCGGGT. Trimmed reads were aligned to the reference genome using STARSolo (version 2.7.3a_2020-05-22) with a 50-base overhang (--sjdbOverhang 50) with the following parameters: --seedSearchStartLmax 10 –alignIntronMax 1000000 --outFilterScoreMin 0 --outFilterMultimapNmax 1 --chimScoreSeparation 10 --chimScoreMin 20 --chimSegmentMin 15 --outFilterMismatchNmax 5. Aligned reads were deduplicated with UMI-tools (version 1.0.1) using --spliced-is-unique --per-cell --read-length --no-sort-output, and sorted using sambamba (version 0.7.1). Finally, aligned and deduplicated reads were parsed to extract the mapping coordinates and the reference sequence contexts around both the 5’ and 3’ ends of the reads.
Count tables report the number of unique RPFs aligning within any annotated coding sequence for each gene. Reads were only counted if they aligned to the correct strand, if the trimmed length was between 30 and 45 nt long, and if the trimmed length was equal to the mapped length. Additionally, for these counts, the coding sequence was expanded to include 25 nt upstream and downstream of the start and stop codons, respectively.
A random forest model was trained to predict the A-site location within an RPF read based on the footprint length and the sequence context around the 5’ and 3’ ends. The model was implemented in R (version 3.6.3) using ranger (version 0.12.1) with mlr (version 2.17.1) wrappers for training, tuning, assessment, and prediction. The model was trained on reads spanning a stop codon that satisfied the counting requirements listed above. The number of nucleotides between the 5’ end of these reads and the annotated stop codon was used for training.
hTERT-RPE1-FUCCI cells were only included in downstream analyses if they had more than 4500 unique reads aligning to protein-coding genes, and if more than 85% aligned to coding sequences.
Supplementary_files_format_and_content: Metadata file: Cell-level metadata for cells passing QC. Column Headers: CB: Cell barcode. treatment: Culture/starvation condition prior to sequencing. well: Well location in plate. plate: Plate name. cds: Number of unique reads aligning to coding sequences. utr5: Number of unique reads aligning to 5' UTRs. utr3: Number of unique reads aligning to 3' UTRs. cell_total: Total number of unique reads aligning to protein-coding genes. cds_frac: Fraction of unique total reads aligning to CDS.
Supplementary_files_format_and_content: Count file: Cell-level count tables denoting the number of unique reads aligned to CDS regions for each gene.
Supplementary_files_format_and_content: Plate layout file: Description of the plate layout. Column Headers: well: well location in plate. CB: cell barcode for this well. fraction: Cell fraction/population/treatment.
Genome_build: hg38

organism: Homo sapiens

source_name: hTERT RPE1 FUCCI cells from either interphase or contact-inhibition G0 fractions.

growth_protocol: RPE1 hTERT FUCCI cells (doi: 10.1073/pnas.1322021111) were cultured in DMEM (Gibco) supplemented with 10% FBS (Gibco), 1× GlutaMAX (Gibco) and 1× Pen-Strep (Gibco) at 37 °C 5% CO2.

extract_protocol:
Single cells were index sorted using a BD FACS Influx into 384-well hardshell plates (BioRad) that were pre-filled with 5 µL of light mineral oil (Sigma Aldrich) and 50 nL of lysis buffer [22 mM Tris-HCl pH 7.5, 16.5 mM MgCl2, 5.5 mM CaCl2, 165 mM NaCl, 1.1 % Triton X-100, 2.2 U/µL RNaseIN Plus (Promega), 0.11 mg/mL Cyclohexamide (Sigma Aldrich)]. After sorting, plates were spun down at 2000 ×g for 2 minutes and kept on wet ice until all plates were ready for further processing. No RNA purification was performed.
To generate ribosome-protected footprints, 50 nL of Micrococcal Nuclease (MNase, 10500 U/µL, New England Biolabs) was added to each well, and plates were incubated at 37 °C for 30 minutes. In order to stop digestion, 50 nL of stop mix [0.0186 U/µL Thermolabile Proteinase K (New England Biolabs), 62 mM EGTA (Sigma Aldrich), 16.5 mM EDTA (Ambion), and 697.5 mM guanidium thiocyanite (GuSCN, Sigma Aldrich)] was added to each well, and plates were incubated at 37 °C for 30 minutes then 55 °C for 10 minutes and held at 4 °C.
After ribosome footprint digestion, libraries were constructed using a one-pot small-RNA library preparation protocol that incorporated end repair, two RNA ligations, cDNA synthesis, and an indexing PCR. First, 50 nL of end-repair mix [4.1× of 10× T4 RNA Ligase Buffer (New England Biolabs), 16.4 mM MgCl2, 4.1 mM uridine triphosphate (New England Biolabs), 1.37 U/µL T4  Polynucleotide Kinase (New England Biolabs), and 0.82 U/µL RNaseIN Plus] was added to each well, and plates were incubated at 37 °C for 1 hour and held at 4 °C. Next, 264 nL of 3’ ligation brew [1× T4 RNA Ligase Buffer (New England Biolabs), 1 µM pre-adenylated 3’ adapter (OMV630 Integrated DNA Technologies), 35.5 % PEG-8000 (New England Biolabs), 0.1 % Tween-20 (Sigma Aldrich), 1 U/µL RNaseIN Plus, and 21.3 U/µL T4 RNA Ligase 2 Truncated KQ (New England Biolabs)] was added to each well and plates were incubated at 4 °C for 18 hours. The cDNA synthesis primer was then pre-annealed to the 3’ ligation products by adding 50 nL of the RT primer mix [5.2 µM RT primer (OMV572, Integrated DNA Technologies), 13.5 µM adenosine triphosphate (ATP, New England Biolabs), and 1 % Tween-20] to each well, heating to 65 °C for 1 minute, 37 °C for 2 minutes, 25 °C for 2 minutes, and holding at 4 °C. Five-prime adapters were then ligated by adding 156 nL of 5’ ligation brew [1× T4 RNA Ligase Buffer, 30.75 % PEG-8000, 0.1 % Tween-20, 0.5 µM 5’ adapter (OMV632, Integrated DNA Technologies), 1.25 U/µL T4 RNA Ligase 1 (Ambion)] and incubating at 37 °C for 2 hours and holding at 4 °C. Complementary DNA synthesis was then peformed by adding 771 nL of reverse transcription brew [1.88× 5× RT Buffer (ThermoFisher Scientific), 1.25 mM dNTPs (Promega), 0.1875 % Tween-20, 1.875 U/µL RNaseIN Plus, and 9.375 U/µL Maxima H Minus Reverse Transcriptase (ThermoFisher Scientific)] to each well, and heating at 50 °C for 1 hour, then 85 °C for 5 minutes and holding at 4 °C. Finally, single-cell libraries were indexed during PCR by first transferring 150 nL of 20 µM unique forward index primers (Integrated DNA Technologies) and 3.2 µL of PCR brew [1.5× Q5 Hot Start High-Fidelity 2× Master Mix (New England Biolabs), 0.15 % Tween-20, and 0.94 µM reverse index primer (Integrated DNA Technologies)] to each well. Plates were then incubated at 98 °C for 30 s followed by 10 cycles of 98 °C for 15 s, 65 °C 30 s, 72 °C for 30 s, and then a final incubation at 72 °C for 5 min and holding at 4 °C. Plates were then frozen at -20 °C until pooling.
After library construction the plates were pooled and purified. The contents of each plate were first collected in VBLOK200 reservoirs (Click Bio) by centrifuging at 2000 ×g for 2 min. The aqueous phase (~1.9 mL per plate) was separated from the light mineral oil by centrifugation, and concentrated to approximately 500 µL using n-butanol (Sigma Aldrich) and diethyl ether (Sigma Aldrich). Product was then cleaned up using AMPure XP beads (Beckman Coulter) that had been diluted 5× in bead binding buffer [20 % PEG-8000 (Sigma Aldrich) 2.5 M Sodium Chloride (Sigma Aldrich)]; diluted beads were added to the sample at a 2.1:1 ratio, and the final product was resuspended in 50 µL of low TE buffer [LoTET, 3 mM Tris-HCl pH 8.0 (Ambion), 0.2 mM EDTA pH 8.0 (Gibco), 0.1 % Tween-20]. Half of each of the cleaned-up library pools was then run on a 10 cm 7 % polyacrylamide gel at 200 V for ~6 h, and the ~10 base-pair region from 175-185, corresponding to an insert size of ~30-40 nt was excised. The band was then crushed and soaked overnight at 4 °C in elution buffer [5:1 LoTET:7.5 M ammonium acetate (Sigma Aldrich)]. Finally, eluate was precipitated in ethanol.
adapters: barcode: CAGATC; 5' Adapter: rCrUrArCrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrUr(N1:25252525)r(N1)r(N1)r(N1)r(N1)r(N1)r(N1)r(N1)r(N1)r(N1); 3' Adapter: /5rApp/TGGAATTCTCGGGTGCCAAGG/3ddC/

treatment_protocol: For the interphase fraction, 7.5E4 cells were plated in a MW-6 and collected by trypsinization (TrypLE, Gibco) 36 hours later. For the G0-arrested fraction, 1E5 cells were plated in a MW-24 and collected 72 hours later by trypsinization. DAPI (ThermoFisher Scientific) was added to cultures as a viability marker, and only viable cells were sorted.

tag: cell type

value: hTERT RPE-1

tag: tissue lineage

value: Retina, eye; pigmented epithelium

tag: genetic tag

value: Fluorescent Ubiquitination-based Cell-Cycle Indicators (FUCCI)

tag: cell fraction

value: Contact-inhibition G0 and interphase cells. Specific cell fraction for each well is defined in RPE_G0_layout.csv.gz.

tag: treatment

value: Interphase and G0 treatments


title: HEK293T

description:
processed_data_file: HEK293T_metadata.csv.gz
processed_data_file: HEK293T_RPFcounts.csv.gz
processed_data_file: HEK293T_layout.csv.gz

data_processing:
See github repository https://github.com/mvanins/scRiboSeq_manuscript for code and detailed instructions.
Basecalls were performed on the Illumina NextSeq 500. Reads from each plate were demultiplexed using bcl2fastq (version 2.20.0.422) with --use-bases-mask Y*,I*,Y* --no-lane-splitting --mask-short-adapter-reads 0 --minimum-trimmed-read-length 0. Read 1 contains the insert and Read 2 contains the cell barcode.
Next, the UMI was extracted from the first 10 bases of read 1 and concatenated to the start of the cell barcode. Adapter sequences were then trimmed from read 1 using cutadapt (version 2.7) with -m 15: -a TGGAATTCTCGGGT. Trimmed reads were aligned to the reference genome using STARSolo (version 2.7.3a_2020-05-22) with a 50-base overhang (--sjdbOverhang 50) with the following parameters: --seedSearchStartLmax 10 –alignIntronMax 1000000 --outFilterScoreMin 0 --outFilterMultimapNmax 1 --chimScoreSeparation 10 --chimScoreMin 20 --chimSegmentMin 15 --outFilterMismatchNmax 5. Aligned reads were deduplicated with UMI-tools (version 1.0.1) using --spliced-is-unique --per-cell --read-length --no-sort-output, and sorted using sambamba (version 0.7.1). Finally, aligned and deduplicated reads were parsed to extract the mapping coordinates and the reference sequence contexts around both the 5’ and 3’ ends of the reads.
Count tables report the number of unique RPFs aligning within any annotated coding sequence for each gene. Reads were only counted if they aligned to the correct strand, if the trimmed length was between 30 and 45 nt long, and if the trimmed length was equal to the mapped length. Additionally, for these counts, the coding sequence was expanded to include 25 nt upstream and downstream of the start and stop codons, respectively.
A random forest model was trained to predict the A-site location within an RPF read based on the footprint length and the sequence context around the 5’ and 3’ ends. The model was implemented in R (version 3.6.3) using ranger (version 0.12.1) with mlr (version 2.17.1) wrappers for training, tuning, assessment, and prediction. The model was trained on reads spanning a stop codon that satisfied the counting requirements listed above. The number of nucleotides between the 5’ end of these reads and the annotated stop codon was used for training.
HEK293T cells were only included in downstream analyses if they had more than 1000 unique reads aligning to protein-coding genes, and if more than 75% aligned to coding sequences.
Supplementary_files_format_and_content: Metadata file: Cell-level metadata for cells passing QC. Column Headers: CB: Cell barcode. treatment: Culture/starvation condition prior to sequencing. well: Well location in plate. plate: Plate name. cds: Number of unique reads aligning to coding sequences. utr5: Number of unique reads aligning to 5' UTRs. utr3: Number of unique reads aligning to 3' UTRs. cell_total: Total number of unique reads aligning to protein-coding genes. cds_frac: Fraction of unique total reads aligning to CDS.
Supplementary_files_format_and_content: Count file: Cell-level count tables denoting the number of unique reads aligned to CDS regions for each gene.
Supplementary_files_format_and_content: Plate layout file: Description of the plate layout. Column Headers: well: well location in plate. CB: cell barcode for this well. fraction: Cell fraction/population/treatment.
Genome_build: hg38

source_name: HEK293T cells that were cultured in DMEM + 10% FBS

growth_protocol: HEK293T cells were cultured in rich DMEM medium based on powdered DMEM for SILAC (ThermoFisher Scientific) supplemented with 10% dialized FBS (ThermoFisher Scientific), 1× GlutaMAX (Gibco), and 1× Pen-Strep (Gibco) at 37 °C and 5% CO2.

extract_protocol:
Single cells were sorted using a BD FACS Influx into 384-well hardshell plates (BioRad) that were pre-filled with 5 µL of light mineral oil (Sigma Aldrich) and 50 nL of lysis buffer [22 mM Tris-HCl pH 7.5, 16.5 mM MgCl2, 5.5 mM CaCl2, 165 mM NaCl, 1.1 % Triton X-100, 2.2 U/µL RNaseIN Plus (Promega), 0.11 mg/mL Cyclohexamide (Sigma Aldrich)]. After sorting, plates were spun down at 2000 ×g for 2 minutes and kept on wet ice until all plates were ready for further processing. No RNA purification was performed.
To generate ribosome-protected footprints, 50 nL of Micrococcal Nuclease (MNase, 10500 U/µL, New England Biolabs) was added to each well, and plates were incubated at 37 °C for 30 minutes. In order to stop digestion, 50 nL of stop mix [0.0186 U/µL Thermolabile Proteinase K (New England Biolabs), 62 mM EGTA (Sigma Aldrich), 16.5 mM EDTA (Ambion), and 697.5 mM guanidium thiocyanite (GuSCN, Sigma Aldrich)] was added to each well, and plates were incubated at 37 °C for 30 minutes then 55 °C for 10 minutes and held at 4 °C.
After ribosome footprint digestion, libraries were constructed using a one-pot small-RNA library preparation protocol that incorporated end repair, two RNA ligations, cDNA synthesis, and an indexing PCR. First, 50 nL of end-repair mix [4.1× of 10× T4 RNA Ligase Buffer (New England Biolabs), 16.4 mM MgCl2, 4.1 mM uridine triphosphate (New England Biolabs), 1.37 U/µL T4  Polynucleotide Kinase (New England Biolabs), and 0.82 U/µL RNaseIN Plus] was added to each well, and plates were incubated at 37 °C for 1 hour and held at 4 °C. Next, 264 nL of 3’ ligation brew [1× T4 RNA Ligase Buffer (New England Biolabs), 1 µM pre-adenylated 3’ adapter (OMV630 Integrated DNA Technologies), 35.5 % PEG-8000 (New England Biolabs), 0.1 % Tween-20 (Sigma Aldrich), 1 U/µL RNaseIN Plus, and 21.3 U/µL T4 RNA Ligase 2 Truncated KQ (New England Biolabs)] was added to each well and plates were incubated at 4 °C for 18 hours. The cDNA synthesis primer was then pre-annealed to the 3’ ligation products by adding 50 nL of the RT primer mix [5.2 µM RT primer (OMV572, Integrated DNA Technologies), 13.5 µM adenosine triphosphate (ATP, New England Biolabs), and 1 % Tween-20] to each well, heating to 65 °C for 1 minute, 37 °C for 2 minutes, 25 °C for 2 minutes, and holding at 4 °C. Five-prime adapters were then ligated by adding 156 nL of 5’ ligation brew [1× T4 RNA Ligase Buffer, 30.75 % PEG-8000, 0.1 % Tween-20, 0.5 µM 5’ adapter (OMV632, Integrated DNA Technologies), 1.25 U/µL T4 RNA Ligase 1 (Ambion)] and incubating at 37 °C for 2 hours and holding at 4 °C. Complementary DNA synthesis was then peformed by adding 771 nL of reverse transcription brew [1.88× 5× RT Buffer (ThermoFisher Scientific), 1.25 mM dNTPs (Promega), 0.1875 % Tween-20, 1.875 U/µL RNaseIN Plus, and 9.375 U/µL Maxima H Minus Reverse Transcriptase (ThermoFisher Scientific)] to each well, and heating at 50 °C for 1 hour, then 85 °C for 5 minutes and holding at 4 °C. Finally, single-cell libraries were indexed during PCR by first transferring 150 nL of 20 µM unique forward index primers (Integrated DNA Technologies) and 3.2 µL of PCR brew [1.5× Q5 Hot Start High-Fidelity 2× Master Mix (New England Biolabs), 0.15 % Tween-20, and 0.94 µM reverse index primer (Integrated DNA Technologies)] to each well. Plates were then incubated at 98 °C for 30 s followed by 10 cycles of 98 °C for 15 s, 65 °C 30 s, 72 °C for 30 s, and then a final incubation at 72 °C for 5 min and holding at 4 °C. Plates were then frozen at -20 °C until pooling.
After library construction the plates were pooled and purified. The contents of each plate were first collected in VBLOK200 reservoirs (Click Bio) by centrifuging at 2000 ×g for 2 min. The aqueous phase (~1.9 mL per plate) was separated from the light mineral oil by centrifugation, and concentrated to approximately 500 µL using n-butanol (Sigma Aldrich) and diethyl ether (Sigma Aldrich). Product was then cleaned up using AMPure XP beads (Beckman Coulter) that had been diluted 5× in bead binding buffer [20 % PEG-8000 (Sigma Aldrich) 2.5 M Sodium Chloride (Sigma Aldrich)]; diluted beads were added to the sample at a 2.1:1 ratio, and the final product was resuspended in 50 µL of low TE buffer [LoTET, 3 mM Tris-HCl pH 8.0 (Ambion), 0.2 mM EDTA pH 8.0 (Gibco), 0.1 % Tween-20]. Half of each of the cleaned-up library pools was then run on a 10 cm 7 % polyacrylamide gel at 200 V for ~6 h, and the ~10 base-pair region from 175-185, corresponding to an insert size of ~30-40 nt was excised. The band was then crushed and soaked overnight at 4 °C in elution buffer [5:1 LoTET:7.5 M ammonium acetate (Sigma Aldrich)]. Finally, eluate was precipitated in ethanol.
adapters: barcode: AGTTCC; 5' Adapter: rCrUrArCrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrUr(N1:25252525)r(N1)r(N1)r(N1)r(N1)r(N1)r(N1)r(N1)r(N1)r(N1); 3' Adapter: /5rApp/TGGAATTCTCGGGTGCCAAGG/3ddC/

treatment_protocol: Cells were cultured to ~70% confluency in rich DMEM (Gibco). Cells were dissociated to a single-cell suspension using TrypLE (Gibco). DAPI (ThermoFisher Scientific) was added to samples as a viability marker and only viable cells were sorted.

value: embryonic kidney

tag: treatment medium

value: DMEM + 10% FBS


title: A4P2

city: beijing

first: shuangqing

last: liu

address: datun street

country: China

institute: institute of biophysics

data_processing:
Illumina HiSeq 2000 software used for basecalling
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to mm10 whole genome using hisat2 version 2.1.0 with parameters --dta-cufflinks
FPKM were calculated using cuffquant following by cuffnorm
Genome_build: mm10
Supplementary_files_format_and_content: excel files include  counts and FPKM values for each Sample

source_name: S100A4+ TAMs

growth_protocol: Murine TAMs that isolated from the E0771 breast cancer cell-bearing S100A4WT-EGFP mice were sorted by flow cytometry into two subset populations, CD45+F4/80+CD11b+GFP+ and CD45+F4/80+CD11b+GFP-.

extract_protocol:
Total RNA was extracted by commerical RNA extraction Kit
A total amount of 1 μg RNA per sample was used as input material for the RNA sample preparations.   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 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, Adaptor with hairpin loop structure were ligated to prepare for hybridization. In order to select cDNA fragments of preferentially 370~420 bp in length, the library fragments were purified with AMPure XP system (Beckman Coulter, Beverly, USA). 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: untreated

tag: tissue

value: from E0771 tumor of S100A4-EGFP mice


title: B16-F10.dmso.A


title: A4P1


title: PSK2: Planktonic-free_Rep2


title: HEK293T-Starv2

description:
processed_data_file: HEK293T_Starv_metadata.csv.gz
processed_data_file: HEK293T_Starv_RPFcounts.csv.gz
processed_data_file: HEK293T_Starv_layout.csv.gz

source_name: HEK293T cells that were cultured in DMEM that was either complete, or that had been depleted of Arginine or Leucine for 3 or 6 hours.

extract_protocol:
Single cells were sorted using a BD FACS Influx into 384-well hardshell plates (BioRad) that were pre-filled with 5 µL of light mineral oil (Sigma Aldrich) and 50 nL of lysis buffer [22 mM Tris-HCl pH 7.5, 16.5 mM MgCl2, 5.5 mM CaCl2, 165 mM NaCl, 1.1 % Triton X-100, 2.2 U/µL RNaseIN Plus (Promega), 0.11 mg/mL Cyclohexamide (Sigma Aldrich)]. After sorting, plates were spun down at 2000 ×g for 2 minutes and kept on wet ice until all plates were ready for further processing. No RNA purification was performed.
To generate ribosome-protected footprints, 50 nL of Micrococcal Nuclease (MNase, 10500 U/µL, New England Biolabs) was added to each well, and plates were incubated at 37 °C for 30 minutes. In order to stop digestion, 50 nL of stop mix [0.0186 U/µL Thermolabile Proteinase K (New England Biolabs), 62 mM EGTA (Sigma Aldrich), 16.5 mM EDTA (Ambion), and 697.5 mM guanidium thiocyanite (GuSCN, Sigma Aldrich)] was added to each well, and plates were incubated at 37 °C for 30 minutes then 55 °C for 10 minutes and held at 4 °C.
After ribosome footprint digestion, libraries were constructed using a one-pot small-RNA library preparation protocol that incorporated end repair, two RNA ligations, cDNA synthesis, and an indexing PCR. First, 50 nL of end-repair mix [4.1× of 10× T4 RNA Ligase Buffer (New England Biolabs), 16.4 mM MgCl2, 4.1 mM uridine triphosphate (New England Biolabs), 1.37 U/µL T4  Polynucleotide Kinase (New England Biolabs), and 0.82 U/µL RNaseIN Plus] was added to each well, and plates were incubated at 37 °C for 1 hour and held at 4 °C. Next, 264 nL of 3’ ligation brew [1× T4 RNA Ligase Buffer (New England Biolabs), 1 µM pre-adenylated 3’ adapter (OMV630 Integrated DNA Technologies), 35.5 % PEG-8000 (New England Biolabs), 0.1 % Tween-20 (Sigma Aldrich), 1 U/µL RNaseIN Plus, and 21.3 U/µL T4 RNA Ligase 2 Truncated KQ (New England Biolabs)] was added to each well and plates were incubated at 4 °C for 18 hours. The cDNA synthesis primer was then pre-annealed to the 3’ ligation products by adding 50 nL of the RT primer mix [5.2 µM RT primer (OMV572, Integrated DNA Technologies), 13.5 µM adenosine triphosphate (ATP, New England Biolabs), and 1 % Tween-20] to each well, heating to 65 °C for 1 minute, 37 °C for 2 minutes, 25 °C for 2 minutes, and holding at 4 °C. Five-prime adapters were then ligated by adding 156 nL of 5’ ligation brew [1× T4 RNA Ligase Buffer, 30.75 % PEG-8000, 0.1 % Tween-20, 0.5 µM 5’ adapter (OMV632, Integrated DNA Technologies), 1.25 U/µL T4 RNA Ligase 1 (Ambion)] and incubating at 37 °C for 2 hours and holding at 4 °C. Complementary DNA synthesis was then peformed by adding 771 nL of reverse transcription brew [1.88× 5× RT Buffer (ThermoFisher Scientific), 1.25 mM dNTPs (Promega), 0.1875 % Tween-20, 1.875 U/µL RNaseIN Plus, and 9.375 U/µL Maxima H Minus Reverse Transcriptase (ThermoFisher Scientific)] to each well, and heating at 50 °C for 1 hour, then 85 °C for 5 minutes and holding at 4 °C. Finally, single-cell libraries were indexed during PCR by first transferring 150 nL of 20 µM unique forward index primers (Integrated DNA Technologies) and 3.2 µL of PCR brew [1.5× Q5 Hot Start High-Fidelity 2× Master Mix (New England Biolabs), 0.15 % Tween-20, and 0.94 µM reverse index primer (Integrated DNA Technologies)] to each well. Plates were then incubated at 98 °C for 30 s followed by 10 cycles of 98 °C for 15 s, 65 °C 30 s, 72 °C for 30 s, and then a final incubation at 72 °C for 5 min and holding at 4 °C. Plates were then frozen at -20 °C until pooling.
After library construction the plates were pooled and purified. The contents of each plate were first collected in VBLOK200 reservoirs (Click Bio) by centrifuging at 2000 ×g for 2 min. The aqueous phase (~1.9 mL per plate) was separated from the light mineral oil by centrifugation, and concentrated to approximately 500 µL using n-butanol (Sigma Aldrich) and diethyl ether (Sigma Aldrich). Product was then cleaned up using AMPure XP beads (Beckman Coulter) that had been diluted 5× in bead binding buffer [20 % PEG-8000 (Sigma Aldrich) 2.5 M Sodium Chloride (Sigma Aldrich)]; diluted beads were added to the sample at a 2.1:1 ratio, and the final product was resuspended in 50 µL of low TE buffer [LoTET, 3 mM Tris-HCl pH 8.0 (Ambion), 0.2 mM EDTA pH 8.0 (Gibco), 0.1 % Tween-20]. Half of each of the cleaned-up library pools was then run on a 10 cm 7 % polyacrylamide gel at 200 V for ~6 h, and the ~10 base-pair region from 175-185, corresponding to an insert size of ~30-40 nt was excised. The band was then crushed and soaked overnight at 4 °C in elution buffer [5:1 LoTET:7.5 M ammonium acetate (Sigma Aldrich)]. Finally, eluate was precipitated in ethanol.
adapters: barcode: TAATCG; 5' Adapter: rCrUrArCrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrUr(N1:25252525)r(N1)r(N1)r(N1)r(N1)r(N1)r(N1)r(N1)r(N1)r(N1); 3' Adapter: /5rApp/TGGAATTCTCGGGTGCCAAGG/3ddC/

treatment_protocol: Cells were cultured to ~70% confluency in rich DMEM medium based on powdered DMEM for SILAC (ThermoFisher Scientific) that was supplemented with 10% dialized FBS (ThermoFisher Scientific), 105 mg/L L-leucine (Sigma Aldrich), 84 mg/L L-arginine HCl (Sigma Aldrich), and 146 mg/L L-lysine HCl (Sigma Aldrich). Three and six hours before sorting, cells were washed once with phosphate buffered saline (PBS) and resuspended in medium that did not contain either arginine or lysine. Before sorting, cells were mechanically dissociated to a single-cell suspension by pipetting up and down. DAPI (ThermoFisher Scientific) was added to cultures as a viability marker, and only viable cells were sorted.

value: DMEM + 10% Dialized FBS, DMEM + 10% Dialized FBS -Arginine, DMEM + 10% Dialized FBS -Leucine. Specific treatment for each well is defined in HEK293T_Starv_layout.csv.gz.

tag: treatment length

value: 0, 3, and 6 hours. Specific treatment for each well is defined in HEK293T_Starv_layout.csv.gz.


title: PSD1: Planktonic-shedding_Rep1

source_name: Planktonic-shedding cell

value: 24-hr Planktonic-shedding cell growing in Luria-Bertani broth (LB)


title: Colon26 cells from syngeneic mouse model CXD101 treatment replicate 3

city: St. Petersburg

first: Alexander

last: Kanapin

address: 7/9 University Emb.

department: Institute for Translational Biomedicine

country: Russia

institute: St Petersburg Universoty

data_processing:
FASTQ files were trimmed to remove adapters and low-quality bases with TrimGalore v0.4.3
The trimmed reads were aligned to the human and mouse reference genomes (builds hg19 and mm10, respectively) with STAR aligner v.2.7 with two mismatches allowed
Differential gene expression analysis was done with DESeq2 R Bioconductor package (v.1.22), using read counts data provided by the aligner
Genes were considered differentially expressed if the adjusted P-value, calculated using Benjamini-Hochberg method was less than 0.01.
We further filtered significantly differentially expressed gene sets to select for genes expressed at high levels using 2-fold change in absolute expression levels for both human and mouse datasets.
Genome_build: hg19 for human and mm10 for mouse
Supplementary_files_format_and_content: raw counts

source_name: Colon26 cells

growth_protocol: SW620 and CT26 cell lines were grown in high glucose Dulbecco’s modified Eagle medium supplemented with 10% (v/v) FBS and 1% penicillin/streptomycin.

extract_protocol:
total RNA was extracted using Promega ReliaPrep™ RNA Cell Miniprep System
cDNA libraries were made using NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina. SW620 RNAseq was perfomed with Illumina HiSeqTM4000. CT26 and Colon26 RNAseq were perfomed with  BGISeq500.

treatment_protocol: HDAC inhibition was induced with 1uM and 2.7uM CXD101 for 72h in SW620 and CT26 cells, respectvely. Treatment of mouse Colon26 syngeneic  model was conducted with 50mg/kg of CXD101 for 14 days (schema: 5 days on/2 days off).

value: Colorectal carcinoma

tag: conditions

value: HDAC inhibitor treatment:50mg/kg, 14 days, schema 5days on/2 days off


title: Normal4_Expr_norm

value: Sorted human lung mesenchymal cells


title: HEK293T-Starv1

extract_protocol:
Single cells were sorted using a BD FACS Influx into 384-well hardshell plates (BioRad) that were pre-filled with 5 µL of light mineral oil (Sigma Aldrich) and 50 nL of lysis buffer [22 mM Tris-HCl pH 7.5, 16.5 mM MgCl2, 5.5 mM CaCl2, 165 mM NaCl, 1.1 % Triton X-100, 2.2 U/µL RNaseIN Plus (Promega), 0.11 mg/mL Cyclohexamide (Sigma Aldrich)]. After sorting, plates were spun down at 2000 ×g for 2 minutes and kept on wet ice until all plates were ready for further processing. No RNA purification was performed.
To generate ribosome-protected footprints, 50 nL of Micrococcal Nuclease (MNase, 10500 U/µL, New England Biolabs) was added to each well, and plates were incubated at 37 °C for 30 minutes. In order to stop digestion, 50 nL of stop mix [0.0186 U/µL Thermolabile Proteinase K (New England Biolabs), 62 mM EGTA (Sigma Aldrich), 16.5 mM EDTA (Ambion), and 697.5 mM guanidium thiocyanite (GuSCN, Sigma Aldrich)] was added to each well, and plates were incubated at 37 °C for 30 minutes then 55 °C for 10 minutes and held at 4 °C.
After ribosome footprint digestion, libraries were constructed using a one-pot small-RNA library preparation protocol that incorporated end repair, two RNA ligations, cDNA synthesis, and an indexing PCR. First, 50 nL of end-repair mix [4.1× of 10× T4 RNA Ligase Buffer (New England Biolabs), 16.4 mM MgCl2, 4.1 mM uridine triphosphate (New England Biolabs), 1.37 U/µL T4  Polynucleotide Kinase (New England Biolabs), and 0.82 U/µL RNaseIN Plus] was added to each well, and plates were incubated at 37 °C for 1 hour and held at 4 °C. Next, 264 nL of 3’ ligation brew [1× T4 RNA Ligase Buffer (New England Biolabs), 1 µM pre-adenylated 3’ adapter (OMV630 Integrated DNA Technologies), 35.5 % PEG-8000 (New England Biolabs), 0.1 % Tween-20 (Sigma Aldrich), 1 U/µL RNaseIN Plus, and 21.3 U/µL T4 RNA Ligase 2 Truncated KQ (New England Biolabs)] was added to each well and plates were incubated at 4 °C for 18 hours. The cDNA synthesis primer was then pre-annealed to the 3’ ligation products by adding 50 nL of the RT primer mix [5.2 µM RT primer (OMV572, Integrated DNA Technologies), 13.5 µM adenosine triphosphate (ATP, New England Biolabs), and 1 % Tween-20] to each well, heating to 65 °C for 1 minute, 37 °C for 2 minutes, 25 °C for 2 minutes, and holding at 4 °C. Five-prime adapters were then ligated by adding 156 nL of 5’ ligation brew [1× T4 RNA Ligase Buffer, 30.75 % PEG-8000, 0.1 % Tween-20, 0.5 µM 5’ adapter (OMV632, Integrated DNA Technologies), 1.25 U/µL T4 RNA Ligase 1 (Ambion)] and incubating at 37 °C for 2 hours and holding at 4 °C. Complementary DNA synthesis was then peformed by adding 771 nL of reverse transcription brew [1.88× 5× RT Buffer (ThermoFisher Scientific), 1.25 mM dNTPs (Promega), 0.1875 % Tween-20, 1.875 U/µL RNaseIN Plus, and 9.375 U/µL Maxima H Minus Reverse Transcriptase (ThermoFisher Scientific)] to each well, and heating at 50 °C for 1 hour, then 85 °C for 5 minutes and holding at 4 °C. Finally, single-cell libraries were indexed during PCR by first transferring 150 nL of 20 µM unique forward index primers (Integrated DNA Technologies) and 3.2 µL of PCR brew [1.5× Q5 Hot Start High-Fidelity 2× Master Mix (New England Biolabs), 0.15 % Tween-20, and 0.94 µM reverse index primer (Integrated DNA Technologies)] to each well. Plates were then incubated at 98 °C for 30 s followed by 10 cycles of 98 °C for 15 s, 65 °C 30 s, 72 °C for 30 s, and then a final incubation at 72 °C for 5 min and holding at 4 °C. Plates were then frozen at -20 °C until pooling.
After library construction the plates were pooled and purified. The contents of each plate were first collected in VBLOK200 reservoirs (Click Bio) by centrifuging at 2000 ×g for 2 min. The aqueous phase (~1.9 mL per plate) was separated from the light mineral oil by centrifugation, and concentrated to approximately 500 µL using n-butanol (Sigma Aldrich) and diethyl ether (Sigma Aldrich). Product was then cleaned up using AMPure XP beads (Beckman Coulter) that had been diluted 5× in bead binding buffer [20 % PEG-8000 (Sigma Aldrich) 2.5 M Sodium Chloride (Sigma Aldrich)]; diluted beads were added to the sample at a 2.1:1 ratio, and the final product was resuspended in 50 µL of low TE buffer [LoTET, 3 mM Tris-HCl pH 8.0 (Ambion), 0.2 mM EDTA pH 8.0 (Gibco), 0.1 % Tween-20]. Half of each of the cleaned-up library pools was then run on a 10 cm 7 % polyacrylamide gel at 200 V for ~6 h, and the ~10 base-pair region from 175-185, corresponding to an insert size of ~30-40 nt was excised. The band was then crushed and soaked overnight at 4 °C in elution buffer [5:1 LoTET:7.5 M ammonium acetate (Sigma Aldrich)]. Finally, eluate was precipitated in ethanol.
adapters: barcode: CTCAGA; 5' Adapter: rCrUrArCrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrUr(N1:25252525)r(N1)r(N1)r(N1)r(N1)r(N1)r(N1)r(N1)r(N1)r(N1); 3' Adapter: /5rApp/TGGAATTCTCGGGTGCCAAGG/3ddC/


title: Normal2_Expr_norm

value: Healthy


title: Colon26 cells from syngeneic mouse model CXD101 treatment replicate 4


title: Colon26 cells from syngeneic mouse model CXD101 treatment replicate 2


title: Colon26 cells from syngeneic mouse model CXD101 treatment replicate 1


title: A4N3

source_name: S100A4- TAMs


title: Normal1_Expr_norm

value: scRNA-sequencing


title: IPF5_Expr_norm

source_name: IPF human lung

value: IPF


title: Colon26 cells from syngeneic mouse model replicate 3

value: no treatment


title: A4N2


title: Normal6_Expr_norm


title: A4P3


title: A4N1


title: Colon26 cells from syngeneic mouse model replicate 2


title: IPF4_Expr_norm


title: NIC AXL KO hypoxia rep 3

city: Boston

first: Marie-Anne

last: Goyette

address: 450 Brookline avenue

institute: DFCI

data_processing:
Quality of the raw reads was assessed with FastQC 0.11.5
Reads were aligned to the GRCm38 genome with Star 2.5.1b
Raw alignment counts were calculated with featureCounts 1.4.6
Differential expression measurements were performed with DESeq2 1.12.3
Genome_build: GRCm38
Supplementary_files_format_and_content: tab-delimited text file with raw alignment counts of each sample
Supplementary_files_format_and_content: excel file of Deseq analysis including raw counts normalized relative to the total number of reads

source_name: MMTV-NIC breast cancer cells

extract_protocol:
Total RNA was extracted using RNeasy columns (Qiagen).
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: treament

tag: genotype

value: AXL KO


title: IPF3_Expr_raw


title: Colon26 cells from syngeneic mouse model replicate 1


title: NIC AXL KO hypoxia rep 2


title: IPF2_Expr_norm


title: NIC AXL KO hypoxia rep 1


title: CT26 CXD101 cell line RNA-seq replicate 3

source_name: CT26 stable cells

value: HDAC inhibitor treatment:2.7uM, 72h


title: CT26 CXD101 cell line RNA-seq replicate 2


title: IPF1_Expr_norm


title: CT26 CDX101 cell line RNA-seq replicate 1


title: 422: TG sample 21

city: Memphis

first: Canan

last: Kuscu

address: 71 S Manassas St.

institute: University of Tennessee Health Science Center

description: small RNA library sequencing

data_processing:
For small RNA-seq, unitas (version unitas_1.7.0) was used to cut adaptor AGATCGGAAGAG and reads smaller than 18 were filtered out.
To quantify small RNAs, unitas v1.7.0 (with SeqMap v1.0.12 ) was used to map the reads to human sequence of miRBase Release 21, genomic tRNA database, Ensembl Release 88 and SILVA rRNA database Release 128.
edgeR was used on count matrix of tRFs and miRNAs. filterByExpr function of edgeR is used to keep rows that have worthwhile counts in a minimum number of samples. The calcNormFactors function of edgeR is used to calculate effective library size and to normalize for RNA composition by finding trimmed mean of M-values (TMM) between each pair of samples. Tagwise and common dispersions were estimated using the estimateDisp function.
Supplementary_files_format_and_content: excel sheet with counts for every sRNA category in TG group
Supplementary_files_format_and_content: excel sheet with counts for every sRNA category in normal allograft function group
Supplementary_files_format_and_content: excel sheet for differentially expressed tRNA fragments(output of edgeR) with log fold change, log cpm, P value and FDR
Supplementary_files_format_and_content: excel sheet for differentially expressed miRNAs(output of edgeR) with log fold change, log cpm, P value and FDR

source_name: serum

extract_protocol:
Small RNAs were purified from 200ul serum using Qiagen miRNeasy Serum/Plasma Advanced kit (Cat No. 217204).
sRNA library was made using NEBNext small RNA library Prep Set (Cat No. E7330L) according to the manual.

tag: condition

value: Transplant glomerulopathy (TG)

value: serum

tag: molecule subtype

value: small RNA


title: NIC AXL KO normoxia rep 2

value: AXL WT


title: 244: TG sample 19

description: small RNA library sequencing


title: NIC AXL KO normoxia rep 1


title: CT26 cell line RNA-seq replicate 2


title: 279: TG sample 20

description: small RNA library sequencing


title: NIC WT hypoxia rep 3


title: CT26 cell line RNA-seq replicate 3


title: 238: TG sample 18

description: small RNA library sequencing


title: NIC WT hypoxia rep 2


title: NIC WT hypoxia rep 1


title: NIC WT normoxia rep 2


title: NIC AXL KO normoxia rep 3


title: 155: TG sample 15

description: small RNA library sequencing


title: 153: TG sample 14

description: small RNA library sequencing


title: 176: TG sample 16

description: small RNA library sequencing


title: 151: TG sample 13

description: small RNA library sequencing


title: 143: TG sample 11

description: small RNA library sequencing


title: 149: TG sample 12

description: small RNA library sequencing


title: CT26 cell line RNA-seq replicate 1


title: 85: TG sample 9

description: small RNA library sequencing


title: 231: TG sample 17

description: small RNA library sequencing


title: WAT 6D 5mc-IP

city: Shanghai

first: Wei

last: Qi

state: Shanghai

address: B514, L Building

department: School of Life Science and Technology

institute: ShanghaiTech University

library_source: genomic

data_processing:
the Chip-seq reads were with FastQC for basic run quality metrics
the Chip-seq reads were aligned to the mouse genomeusing the Bowtie2 (v2.3.0) algorithm
Differential methylation regions were calculated by the Bioconductor package 'MEDIP' (the main parameters: win-size=500bp, adj.p.value=0.1)
These regions were annotated with annotatePeaks.pl which comes from HOMER
Genome_build: mm10
Supplementary_files_format_and_content: bw files were generated using …; Scores represent …

molecule: genomic DNA

source_name: C3H10T cells

growth_protocol: All cells were maintained at 37°C in humidified air with 5% CO2. C3H10T1/2  preadipocytes were cultured in Dulbecco's modified Eagle medium (DMEM) supplemented with 10% fetal bovine serum (FBS) (growth medium). Growth medium was replaced every 2 days.

extract_protocol:
Lysates were clarified from sonicated nuclei and DNA complexes were isolated with 5mc or 5hmc antibody.
Sequencing analysis was done by Wuxi biology, Briefly, Sequencing libraries were generated by pooling immunoprecipitated DNA from eight samples. Libraries were amplified using Illumina Enrichment Primers.

treatment_protocol: Cells were cultured for an additional day after reaching confluency, and the growth medium was then replaced (day 0) with DMEM supplemented with 10% FBS, 1 μM dexamethasone, 0.5 mM 3-isobutyl-1-methylxanthine and 10 μg/ml insulin (differentiation medium, MDI), or MDI with 1 μM rosiglitazone and 1 nM T3 (differentiation medium, MDI+RT). After 48 h, the differentiation medium was replaced (day 2) with DMEM + 10% FBS containing 5 μg/ ml insulin, and the cells were allowed to accumulate lipid droplets until experimental use.

tag: chip antibody


title: BAT 6D input

value: input


title: NIC WT normoxia rep 3


title: 72: TG sample 8

description: small RNA library sequencing


title: NIC WT normoxia rep 1


title: WAT 6D 5hmc-IP


title: WAT 6D input


title: 54: TG sample 7

description: small RNA library sequencing


title: BAT 6D 5mc-IP


title: BAT 6D 5hmc-IP


title: WAT 4D input


title: 101: TG sample 10

description: small RNA library sequencing


title: 52: TG sample 6

description: small RNA library sequencing


title: WAT 4D 5hmc-IP


title: WAT 4D 5mc-IP


title: 94RYH63

type: RNA

city: Sheffield

first: Ilaria

last: Giovannelli

address: 385a Glossop Road

department: Sheffield Institute for Translational Neuroscience

country: United Kingdom

institute: University of Sheffield

description: Gene expression data from human peripheral blood

hyb_protocol: Fragmented and labelled ss cDNA was hybridized on Human Clariom D Affymetrix® chips.

scan_protocol: The GeneChips were scanned in the Affymetrix GCS3000 scanner and the resultant image processed using the AGCC software package to prepare the .CEL files

data_processing: Gene expression data was normalised using the signal space transformation robust multi-chip analysis (SST-RMA) method. Data was analysed using TAC and Limma software.

label: Biotin

label_protocol: 200ng of total RNA was used to produce antisense RNA (cRNA) and cDNA which was then fragmented and biotinylated using Affymetrix GeneChip® WT PLUS Kit according to manufacturer's protocol.

extract_protocol:
White blood cell isolation: White blood cells were isolated using LeukoLOCK TM filters
Total RNA extraction: Total RNA was isolated using the LeukoLOCK TM Total RNA isolation kit (AmbionTM) as per manufacturer's instructions.

tag: diagnosis

value: ALS

value: peripheral blood

value: Placebo

tag: time point


title: 96GFO65


title: 63VCW65


title: 63ORT67


title: 45XNH76


title: 33: TG sample 4

description: small RNA library sequencing


title: Urothelial carcinoma, patient B_AAN_27

city: Lyon

first: Jiri

last: Zavadil

address: 150 cours Albert Thomas

country: France

institute: International Agency for Research on Cancer (IARC)

description: Gene expression data from Urothelial carcinoma, patient B_AAN_27

hyb_protocol: 3.75 mcg of biotin-labeled cRNA were hybridized for 16 hrs onto the Affymetrix Human Genome U133 Plus 2.0 Arrays

scan_protocol: Affymetrix 3000 7G GeneArray scanner

data_processing: Feature intensity was extracted by GeneChip Operating System as CEL files. The probe-level analysis of the CEL files was performed by the Robust Multichip Average (RMA) algorithm using the Gene Spring GX10 software (Agilent Technologies, Santa Clara, CA, USA) program. No further adjustments were made to the data in the table.

label: biotin

source_name: Surgically dissected urothelial carcinoma

label_protocol: The RNA was processed using 3' IVT Express kit (Affymetrix), 500 ng of total purified RNA was reverse-transcribed into cDNA and then transcribed into biotin-labelled cRNA.

extract_protocol: Total RNA was isolated from fresh samples of tumor and adjacent normal tissue using using miRNAeasy kit (Qiagen, Valencia, CA, USA) according to the manufacturer’s instructions and  stored in RNAlater. All samples were treated with DNAse (Qiagen, Valencia, CA, USA). RNA samples were stored at −80°C. The quantity and purity of RNA was measured using NanoDrop 2000c (Thermo Scientific, Wilmington, DE, USA). Agilent 2100 Bioanalyser (Agilent Technologies, Santa Clara, CA, USA) was used to determine the RNA integrity number which ranged between 8-10.

tag: individual

value: patient B_AAN_27

tag: gender

value: female

tag: age at surgery

value: 75 years

value: UTUC tumor


title: 49KHX86


title: 53UCM47


title: BAT 4D input

tag: cell type


title: 27: TG sample 3

description: small RNA library sequencing


title: 77PVZ38


title: Urothelial carcinoma, patient B_AAN_25

description: Gene expression data from Urothelial carcinoma, patient B_AAN_25

value: patient B_AAN_25

value: male


title: Urothelial carcinoma, patient B_AAN_21

description: Gene expression data from Urothelial carcinoma, patient B_AAN_21

value: patient B_AAN_21

value: 83 years


title: Urothelial carcinoma, patient B_AAN_22

description: Gene expression data from Urothelial carcinoma, patient B_AAN_22

value: patient B_AAN_22

value: 59 years


title: 49: TG sample 5

description: small RNA library sequencing

value: Transplant glomerulopathy (TG)

value: serum


title: Urothelial carcinoma, patient B_AAN_26

description: Gene expression data from Urothelial carcinoma, patient B_AAN_26

value: patient B_AAN_26

value: 80 years


title: 99WQZ38

value: ALS

value: 2MIU IL2


title: 72EBF68

value: peripheral blood

value: Placebo


title: 36VVR32


title: 44PBR28


title: 44FGS45


title: BAT 4D 5mc-IP


title: 17: TG sample 1

description: small RNA library sequencing

value: small RNA


title: BAT 4D 5hmc-IP


title: 370: Normal sample 9

description: small RNA library sequencing

value: Normal Allograft function


title: wat 2D input

value: input


title: wat 2D 5hmc-IP


title: Urothelial carcinoma, patient B_AAN_18

description: Gene expression data from Urothelial carcinoma, patient B_AAN_18

value: patient B_AAN_18

value: female

value: 76 years

value: UTUC tumor


title: 337: Normal sample 8

description: small RNA library sequencing


title: wat 2D 5mc-IP


title: 323: Normal sample 7

description: small RNA library sequencing


title: BAT 2D input


title: 322: Normal sample 6

description: small RNA library sequencing


title: Urothelial carcinoma, patient B_AAN_20

description: Gene expression data from Urothelial carcinoma, patient B_AAN_20

value: patient B_AAN_20


title: 97MAY94


title: 21: TG sample 2

description: small RNA library sequencing


title: 73XJQ42


title: BAT 2D 5hmc-IP


title: Urothelial carcinoma, patient B_AAN_13

description: Gene expression data from Urothelial carcinoma, patient B_AAN_13

value: patient B_AAN_13

value: 71 years


title: 311: Normal sample 5

description: small RNA library sequencing

value: serum


title: 83VZK65


title: BAT 2D 5mc-IP


title: 271: Normal sample 4

description: small RNA library sequencing


title: Urothelial carcinoma, patient B_AAN_07

description: Gene expression data from Urothelial carcinoma, patient B_AAN_07

value: patient B_AAN_07

value: 73 years


description: Gene expression data from Normal urothelial tissue, patient B_AAN_27

source_name: Normal urothelial tissue from tumor surgery

value: patient B_AAN_27

value: 75 years

value: normal urothelium


title: WT 0D input


title: 246: Normal sample 3

description: small RNA library sequencing


title: 57KHM43

value: ALS

value: peripheral blood

value: 2MIU IL2


title: 59XAQ99


title: Urothelial carcinoma, patient B_AAN_04

description: Gene expression data from Urothelial carcinoma, patient B_AAN_04

value: patient B_AAN_04

value: 74 years


title: Urothelial carcinoma, patient B_AAN_06

description: Gene expression data from Urothelial carcinoma, patient B_AAN_06

value: patient B_AAN_06

value: female

value: 78 years


title: Urothelial carcinoma, patient B_AAN_03

description: Gene expression data from Urothelial carcinoma, patient B_AAN_03

value: patient B_AAN_03

value: male

value: 68 years

value: UTUC tumor


title: WT 0D 5hmc-IP


title: 53HHI65


title: WT 0D 5mc-IP


title: 47NSH95


title: 97: Normal sample 1

description: small RNA library sequencing

value: Normal Allograft function

value: serum

value: small RNA


title: 165: Normal sample 2

description: small RNA library sequencing


title: MCF10A, ∆40p53:WTp53 cDNA insert, 375µM 5FU, Replicate 2 [41_DMSO_rep2_S11]

city: Boulder

first: Dylan

middle: J

last: Taatjes

state: CO

address: Campus Box 596

institute: University of Colorado Boulder

data_processing:
The RNA-seq data was processed using a Nextflow pipeline v1.1 (https://github.com/Dowell-Lab/RNAseq-Flow).
Gene counts were generated using featureCounts(Liao, Smyth, and Shi 2014) and differential gene expression analysis was performed using DESeq2(Anders and Huber 2010).
Duplicate genes were filtered and those with the highest FPKM were kept for analysis.
Genome_build: GRCh38
Supplementary_files_format_and_content: counts.FPKM.txt files were generated using featureCounts and contain read quantification (counts) over anotated genes and can be used to feed into DEseq2 to onbtain information about differential expression

source_name: MCF10A_∆40p53:WTp53 cDNA insert_375µM 5FU

extract_protocol:
Total RNA was isolated using TRizol (Invitrogen), and 1ug of total RNA with a RIN number of ≥ 8 was used for RNA-seq library prep.
Sample was enriched for mRNA using NEBNext Poly(A) mRNA Magnetic Isolation Module (E7490) and library was prepared using NEBNext Ultra II Directional RNA Library Prep Kit from Illumina (E7765).

treatment_protocol: MCF10A genome edited cells were seeded on one 10cm dishes (1x107 cells per dish) for each treatment 24 hours prior to the experiments ( 70% confluency at the time of the experiment). Cells were treated simultaneously with 375μM 5FU or 0.375% DMSO for 20 hours.

tag: cell line

value: mammary gland; breast

value: epithelial

tag: crispr/cas9 insertion

value: 3924bp:∆40p53 cDNA tethjered to full length p53 cDNA followed IRES expressed mCherry followed by SV40 polyA sequnece

tag: crispr/cas9 insertion location

value: 375uM 5FU


title: MCF10A, ∆40p53:WTp53 cDNA insert, 375µM 5FU, Replicate 1 [41_DMSO_rep1_S5]

tag: tissue

tag: treatment


title: MCF10A, ∆40p53:WTp53 cDNA insert, DMSO, Replicate 2 [41_5FU_rep2_S12]

source_name: MCF10A_∆40p53:WTp53 cDNA insert_DMSO


description: Gene expression data from Normal urothelial tissue, patient B_AAN_25

value: patient B_AAN_25

value: 75 years

value: normal urothelium


title: 35NXS97

value: ALS

value: peripheral blood

value: 2MIU IL2


title: 35PXK56


title: MCF10A, ∆40p53:WTp53 cDNA insert, DMSO, Replicate 1 [41_5FU_rep1_S6]


description: Gene expression data from Normal urothelial tissue, patient B_AAN_21

value: patient B_AAN_21

value: female

value: 83 years


description: Gene expression data from Normal urothelial tissue, patient B_AAN_26

value: patient B_AAN_26

value: 80 years


description: Gene expression data from Normal urothelial tissue, patient B_AAN_22

value: patient B_AAN_22

value: male

value: 59 years


title: 24WMJ33

description: Gene expression data from human peripheral blood


title: 94GRV28

value: Placebo


title: Varadan_Seedlings-Root_100

city: Hyderabad

first: C

middle: N

last: Neeraja

state: Telangana

address: Principal Scientist, ICAR-IIRR, Rajendranagar, Hyderabad 500030

department: Crop improvement

country: India

institute: ICAR-Indian Institute of Rice Research

hyb_protocol: Amount of signal from the 3’ probe set to either the mid or 5’ probe sets, a measure of the quality of the RNA hybridized to the chip

scan_protocol: Standard Affymetrix protocol .CEL files obtained for each sample were processed through MAS 5 algorithm

data_processing: Present/Marginal/Absent calls were generated for each probe-set, using MAS 5.0 algorithm recommended by Affymetrix. % Present calls should be studied for each sample and if they are greater than 40%, the sample is considered as qualified for the downstream analysis.

organism: Oryza sativa

source_name: root

label_protocol: Standard Affymetrix protocol

growth_protocol: Hydroponics 14 days old seedling root

extract_protocol: The seeds of two varieties viz, Prasanna amd Varadan were washed with sodium hypochlorite followed by five washes of one min each. The seeds were kept in petri plates lined with moistened filter paper for germination.  After seven days the seedlings were transferred to Yoshida medium with nitrogen for 10 days. Then the seedlings were transferred in Yoshida media with supplemented N and without N to observed the response of genotypes under N supplementation and without N supplementation. Total RNA was isolated from roots in replication

tag: variety

value: Varadan

tag: developmental stage

value: 14 days seedling

value: Supplemented nitrogen

value: root


title: Prasanna_Seedlings-Root_100_2

value: Prasanna


description: Gene expression data from Normal urothelial tissue, patient B_AAN_18

value: patient B_AAN_18

value: 76 years

value: normal urothelium


description: Gene expression data from Normal urothelial tissue, patient B_AAN_15

value: patient B_AAN_15

value: 85 years


title: 88TWF48

value: ALS

value: peripheral blood


title: 83QCF22


title: Varadan_Seedlings-Root_0

value: No nitrogen


title: Varadan_Seedlings-Root_100_2


title: 89ERK87


description: Gene expression data from Normal urothelial tissue, patient B_AAN_14

value: patient B_AAN_14


title: MCF10A, WTp53:WTp53 cDNA insert, 375uM 5FU, Replicate 2 [21_DMSO_rep2_S9]

source_name: MCF10A_WTp53:WTp53 cDNA insert_375µM 5FU

value: mammary gland; breast

value: epithelial

value: 4041bp:full length p53 tethered to full length p53 cDNA followed IRES expressed mCherry followed by SV40 polyA sequnece

value: 375uM 5FU


title: 22DSY33

value: 2MIU IL2


title: MCF10A, WTp53:WTp53 cDNA insert, 375uM 5FU, Replicate 1 [21_DMSO_rep1_S3]

source_name: MCF10A_WTp53:WTp53 cDNA insert_375uM 5FU


title: MCF10A, WTp53:WTp53 cDNA insert, DMSO, Replicate 2 [21_5FU_rep2_S10]

source_name: MCF10A_WTp53:WTp53 cDNA insert_DMSO


description: Gene expression data from Normal urothelial tissue, patient B_AAN_19

value: patient B_AAN_19

value: male

value: 66 years


title: 56UWH39

value: Placebo


title: 92NTM59


title: 48CFV44


title: MCF10A, WTp53:WTp53 cDNA insert, DMSO, Replicate 1 [21_5FU_rep1_S4]


title: MCF10A, WTp53 cDNA insert, 375uM 5FU, Replicate 2 [7_5FU_rep2_S8]

source_name: MCF10A_WTp53 cDNA insert_375uM 5FU

value: 2820bp:full length p53 cDNA followed IRES expressed mCherry followed by SV40 polyA sequnece


title: 12-ShnRelKO_PMG-1

city: Heidelberg

first: Erica

last: Valentini

address: Im Neuenheimer Feld 580

country: Germany

institute: DKFZ

data_processing:
Basecalling was performed with RTA 1.18.64.
Salmon (Patro et al. 2017) version 1.0.0 was used to map the reads to the transcriptome with the options -l A -p 8 --validateMappings
The quantification files were then imported and annotated in R using Tximeta (M. I. Love et al. 2020).
The gene expression analysis and differentially expressed genes detection was performed using DESeq2 (Love, Huber, and Anders 2014).
Genes with less than 5 counts among all samples were filtered out and the data were then normalized using the Variance Stabilizing Transformation (VST, Huber et a. 2002).
Genome_build: ENSEMBL BDGP6.22
Supplementary_files_format_and_content: tab separated values (TSV) file with transcript counts for each sample

organism: Drosophila melanogaster

source_name: Posterior Midgut

growth_protocol: The crosses were made at 18 degree and adult flies were shifted to 29 degree to induce RNAi effect.

extract_protocol:
Intestines were dissected and the Crop, malpighian tubules, hindgut were removed, then posterior part of intestine were isolated and flash frozen on dry ice, and RNA was extracted using Qiagen RNeasy kit. Illumina TruSeq RNA Sample Prep Kit (Cat#FC-122-1001) was used with 1 ug of total RNA for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: Temperature sensitive Gal4/UAS system is used, therefore the Drosophila P0 crosses were made and maintained at 18 degree and adult flies P1 were collected shifted to 29 degree to induce RNAi effect.

tag: strain

value: esgts> Shn-RNAi,RelishE20

value: Intestine

tag: age

value: 7 days after shift to 29 degree

tag: genotype


title: 13-ShnRelKO_PMG-2


title: 8-Shn_PMG-1

value: esgts> Shn-RNAi


title: MCF10A, WTp53 cDNA insert, DMSO, Replicate 1 [7_DMSO_rep1_S1]

source_name: MCF10A_WTp53 cDNA insert_DMSO


title: 10-Shn_PMG-3


title: Prasanna_Seedlings-Root_0

value: Prasanna

value: 14 days seedling

value: root


description: Gene expression data from Normal urothelial tissue, patient B_AAN_03

value: patient B_AAN_03

value: 68 years

value: normal urothelium


title: MCF10A, WTp53 cDNA insert, 375uM 5FU, Replicate 1 [7_5FU_rep1_S2]

value: mammary gland; breast

value: epithelial

value: 375uM 5FU


title: 44IKD34

value: ALS

value: peripheral blood


title: Varadan_Seedlings-Shoot_100_2nd replicate [NMA8]

scan_protocol: Affymetrix standard protocol

data_processing:
.CEL files obtained for each sample were processed through MAS 5 algorithm
Present/Marginal/Absent calls were generated for each probe-set, using MAS 5.0 algorithm recommended by Affymetrix. % Present calls should be studied for each sample and if they are greater than 40%, the sample is considered as qualified for the downstream analysis.

source_name: Hydroponics 14 days old seedling shoot  of genotype Varadan

label_protocol: 3' and 5' probes

extract_protocol: The seeds of two varieties viz, Prasanna amd Varadan were washed with sodium hypochlorite followed by five washes of one min each. The seeds were kept in petri plates lined with moistened filter paper for germination.  After seven days the seedlings were transferred to Yoshida medium with nitrogen for 10 days. Then the seedlings were transferred in Yoshida media with supplemented N and without N to observed the response of genotypes under N supplementation and without N supplementation. Total RNA was isolated from shoots in replication

value: Varadan

tag: nitrogen level

value: Supplemented nitrogen

value: 14 days old seedling shoot


description: Gene expression data from Normal urothelial tissue, patient B_AAN_13

value: patient B_AAN_13

value: female

value: 71 years


title: 9-Shn_PMG-2

value: esgts> Shn-RNAi

value: Intestine

value: 7 days after shift to 29 degree


title: Varadan_Seedlings-Shoot_0_1st replicate [NMA5]

source_name: Hydroponics 14 days old seedling shoot of genotype Varadan

value: No nitrogen


title: Prasanna_Seedlings-Root_100

value: Prasanna

value: 14 days seedling

value: root


title: Varadan_Seedlings-Shoot_0_2nd replicate [NMA6]


title: Varadan_Seedlings-Shoot_100_1st replicate [NMA7]


title: 37TMZ28

tag: diagnosis

value: Placebo


title: Prasanna_Seedlings-Shoot_100_1st replicate [NMA3]

source_name: Hydroponics 14 days old seedling shoot of genotype Prasanna


title: 28BMD36

tag: time point


title: 28TBF72


title: MCF10A, WTp53 cDNA insert, DMSO, Replicate 2 [7_DMSO_rep2_S7]

value: mammary gland; breast

value: epithelial

value: 2820bp:full length p53 cDNA followed IRES expressed mCherry followed by SV40 polyA sequnece


title: 5-RelKO_PMG-1

value: esgts> RelishE20


title: 6-RelKO_PMG-2


title: MCF10A, ∆40p53:WTp53 cDNA insert,10uM Nutlin3a, Replicate 1 [RNAseq_41_Nutlin_R1]

description: mRNA (RNA-seq) from MCF10A cells

treatment_protocol: MCF10A genome edited cells were seeded on one 10cm dishes (1x10^7 cells per dish) for each treatment 24 hours prior to the experiments ( 70% confluency at the time of the experiment). Cells were treated simultaneously with 10μM Nutlin3a or 0.1% DMSO for 20 hours.

value: 3924bp:∆40p53 cDNA tethjered to full length p53 cDNA followed IRES expressed mCherry followed by SV40 polyA sequnece

value: 10uM Nutlin3a


title: MCF10A, ∆40p53:WTp53 cDNA insert,10uM Nutlin3a, Replicate 2 [RNAseq_41_Nutlin_R2]

description: mRNA (RNA-seq) from MCF10A cells


title: 24TTF82

value: ALS

value: peripheral blood


title: 88NWQ96

value: 2MIU IL2


title: 72WPD58


title: MCF10A, ∆40p53:WTp53 cDNA insert, DMSO, Replicate 2 [RNAseq_41_DMSO_R2]

description: mRNA (RNA-seq) from MCF10A cells

source_name: MCF10A, ∆40p53:WTp53 cDNA insert, DMSO

value: %0.1 DMSO


title: MCF10A, WTp53:WTp53 cDNA insert,10uM Nutlin3a, Replicate 2 [RNAseq_21_Nutlin_R2]

description: mRNA (RNA-seq) from MCF10A cells

value: 4041bp:full length p53 tethered to full length p53 cDNA followed IRES expressed mCherry followed by SV40 polyA sequnece


title: 6. Shn RNAi PucE69/+

source_name: Midgut

extract_protocol:
Intestines were dissected and the Crop, malpighian tubules, hindgut were removed, then tissues were flash frozen on dry ice, and RNA was extracted using Qiagen RNeasy kit. Illumina TruSeq RNA Sample Prep Kit (Cat#FC-122-1001) was used with 1 ug of total RNA for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: esgts> Shn-RNAi,PucE69/+

value: Intestine

value: 7 days after shift to 29 degree


title: 1-WT_PMG-1

value: esgts> w1118


title: MCF10A, ∆40p53:WTp53 cDNA insert, DMSO, Replicate 1 [RNAseq_41_DMSO_R1]

description: mRNA (RNA-seq) from MCF10A cells


title: Prasanna_Seedlings-Shoot_0_1st replicate [NMA1]

value: Prasanna

value: No nitrogen

value: 14 days old seedling shoot


title: 96HQV85


title: 54SIW45


title: Quercetin-Treated-biological rep2

city: Jeddah

first: Sajjad

last: Karim

state: Makka

address: Sulemania, CEGMR-80216

department: Center of Excellence in Genomic Medicine Research

country: Saudi Arabia

institute: King Abdulaziz University

hyb_protocol: Following fragmentation, 6 µg of cDNA were hybridized for 17 hr at 45C on GeneChip HuGene-1.0-st arrays. GeneChips were washed and stained in the Affymetrix Fluidics Station 400.

scan_protocol: GeneChips were scanned using Affymetrix® GeneChip® Scanner 3000

data_processing: Data were generated using the Affymetrix Expression Console Software using Affymetrix default analysis settings. Data were RMA normalized and analyzed with Partek Genomics Suite 7.0.

source_name: THP-1 treated with Quercetin

label_protocol: Biotinylated cRNAs were prepared according to the standard Affymetrix protocol from 6 µg total RNA (Expression Analysis Technical Manual, 2001, Affymetrix)

growth_protocol: THP-1 monocytes leukemia cells were sub-cultured in T-75 flasks in vertical position. Medium (RPMI) 1640 from (Gibco, USA) were used for this cell line and medium were supplemented with antibiotics (100 μg mL−1 of both streptomycin and penicillin) and (10%) FCS, fetal calf serum.

extract_protocol: Cells were harvested and total RNA was extracted and quality of the purified RNA was verified. RNA concentrations were determined and 250 ng of each RNA sample were processed.

treatment_protocol: THP-1 monocyte were differentiated to macrophage using PMA for 24 hrs, then (THP1) macrophage were treated with DMSO, Punicalagin, Myricetin or Quercetin for 24 hrs.

value: Leukemia Cell Line

tag: passages

value: passage 8

value: Quercetin


title: Myricetin-Treated-biological rep1

source_name: THP-1 treated with Myricetin

value: passage 7

value: Myricetin


title: Quercetin-Treated-biological rep1


title: Prasanna_Seedlings-Shoot_0_2nd replicate [NMA2]


title: Prasanna_Seedlings-Shoot_100_2nd replicate [NMA4]

value: Supplemented nitrogen


title: 7. Shn RNAi PucE69/+


title: 48ZSZ47

value: ALS

value: peripheral blood


title: 3-WT_PMG-3


title: 53WYU63

value: 2MIU IL2


title: MCF10A, WTp53:WTp53 cDNA insert, DMSO, Replicate 1 [RNAseq_21_DMSO_R1]

description: mRNA (RNA-seq) from MCF10A cells

source_name: MCF10A, WTp53:WTp53 cDNA insert, DMSO

value: mammary gland; breast

value: epithelial

value: 4041bp:full length p53 tethered to full length p53 cDNA followed IRES expressed mCherry followed by SV40 polyA sequnece

value: %0.1 DMSO


title: MCF10A, WTp53:WTp53 cDNA insert,10uM Nutlin3a, Replicate 1 [RNAseq_21_Nutlin_R1]

description: mRNA (RNA-seq) from MCF10A cells

value: 10uM Nutlin3a


title: 5. Shn RNAi day7

value: esgts> Shn-RNAi

value: Intestine

value: 7 days after shift to 29 degree


title: MCF10A, WTp53:WTp53 cDNA insert, DMSO, Replicate 2 [RNAseq_21_DMSO_R2]

description: mRNA (RNA-seq) from MCF10A cells


title: 45XVK33


title: 3. WT day7

value: esgts> w1118


title: 4. Shn RNAi day7


title: MCF10A, WTp53 cDNA insert, 10uM Nutlin3a, Replicate 2 [RNAseq_7_Nutlin_R2]

description: mRNA (RNA-seq) from MCF10A cells

value: 2820bp:full length p53 cDNA followed IRES expressed mCherry followed by SV40 polyA sequnece


title: Myricetin-Treated-biological rep2

value: Leukemia Cell Line

value: passage 8


title: 1.WT day7


title: 2. WT day7


title: MCF10A, WTp53 cDNA insert, DMSO, Replicate 1 [RNAseq_7_DMSO_R1]

description: mRNA (RNA-seq) from MCF10A cells

source_name: MCF10A, WTp53 cDNA insert, DMSO


title: MCF10A, WTp53 cDNA insert, 10uM Nutlin3a, Replicate 1 [RNAseq_7_Nutlin_R1]

description: mRNA (RNA-seq) from MCF10A cells


title: MCF10A, ∆40p53:WTp53 cDNA insert, 10uM Nutlin3a, Replicate 3 [41_Nutlin_p53_R3]

description:
DNA bound by p53
processed data file: 41_Nutlin_0.0001_peaks_clean.narrowPeak

data_processing:
All ChIP-seq data was processed (mapped and quality checked) using a Nextflow pipeline, ChIP-Flow v1.0 (https://github.com/Dowell-Lab/ChIP-Flow)
Blacklisted regions (those having artificially high signal and read mapping, http://mitra.stanford.edu/kundaje/akundaje/release/blacklists/hg38-human/) were removed using BEDTools intersect(Quinlan and Hall 2010)
Peak calls were generated using MACS2 narrowPeak. The q-value default cutoff was also decreased from the default of 0.05 to 1e-5.
Genome_build: GRCh38
Supplementary_files_format_and_content: BED6+4 files containing peak locations together with peak summit, p-value, and q-value (.narrowPeak files) called peaks were generated using MACS2 narrowPeak.

extract_protocol:
Six hours after the addition of 10 mM Nutlin-3a or 0.1% DMSO, cells (~60 x 10^6 cells) were fixed with 1% formaldehyde for 10 min at 25oC. Glycine (0.125 M) was then added for 5 min. Cells were scraped into conical tubes and washed with ice cold PBS. Nuclei were isolated by resuspending the cells in NRO buffer (80 ml/million cells; 10 mM Tris-HCl [pH 8], 4 mM MgCl2, 10 mM NaCl, 0.5% [vol/vol] NP-40 and the protease inhibitor cocktail: 0.25 mM phenylmethylsulfonyl fluoride (PMSF), 1 mM sodium metabisulfite, 0.012 TIU/mL aprotinin, 1 mM benzamidine and 1 mM dithiothreitol (DTT)), followed by a 5 min incubation on ice, a low speed spin, and then a final wash with NRO buffer. The isolated nuclei were prepared for shearing based on the Covaris truChIP Chromatin Shearing Kit (Covaris: PN 520237) and sheared for 11 min on Covaris M220 Focused-ultrasonicator. Samples were spun at high speeds then a small aliquot for input was removed while the rest of the supernatant containing the sheared chromatin was transferred to prepared Protein G Dynabeads (Invitrogen: 10003D). 25mL of Protein G Dynabeads beads per pull down were washed 3 times in block solution (PBS, 0.5% BSA) before being resuspended in block solution with 6 mg of DO1 p53 antibody (BD Biosciences: BD554293) per pull down. The beads and antibody were allowed to nutate for 10 min at 25oC before nutating at 4oC for 4 hours. Beads were then washed in block solution once and IP buffer once (15 mM Tris-HCl [pH 8], 150 mM NaCl, 1 mM EDTA, 1% Triton X-100 and the protease inhibitor cocktail). Beads were then resuspended in IP buffer and the sheared chromatin was added. The sample was then allowed to nutate at 4oC for at least 12 hours. The bound chromatin was then washed 3 times with IP buffer, 3 times with RIPA buffer (20 mM Tris-HCl [pH 8], 500 mM NaCl, 1 mM EDTA, 1% Triton X-100 and 0.1% SDS), 2 times with LiCl buffer (20 mM Tris-HCl [pH 8], 500 mM LiCl, 1 mM EDTA, 1% sodium deoxycholate and 1% NP-40) and 2 times with TE Salt buffer (10 mM Tris-HCl [pH 8], 50 mM NaCl, 1 mM EDTA). The sample was then eluted with PK buffer (10 mM Tris-HCl [pH 8], 1 mM EDTA and 1% SDS) containing 100 mg of Proteinase K (New England BioLabs: P81075) per reaction by incubating at 50oC shaking for 1-hour then at 65oC for 1-hour. The supernatant containing the eluted DNA was transferred to a new tube for reverse crosslinking. The input was thawed and brought up to sample volume with PK buffer containing 100 mg of Proteinase K per reaction. All samples and inputs were incubated at 65oC for at least 12 hours. DNA was then purified by phenol chloroform extraction using Light-5PRIME Phase Lock Tubes (Quanta Bio: 2302820) based on the manufacturer’s instructions.
Libraries were prepared using the KAPA Hyper Prep Kit (KAPA Biosystems: KK8502) and sequenced on Illumina NextSeq V2 high output 75-cycle.

treatment_protocol: MCF10A genome edited cells were seeded on six 15cm dishes (1x10^7 cells per dish) for each treatment 24 hours prior to the experiments ( 70% confluency at the time of the experiment). Cells were treated simultaneously with 10μM Nutlin3a or 0.1% DMSO for 6 hour.

value: 3924bp:∆40p53 cDNA tethjered to full length p53 cDNA followed IRES expressed mCherry followed by SV40 polyA sequnece

tag: chip antibody

value: DO-1 (BD Biosciences, BD554293)


title: SLK-kcirc55_4

city: Baltimore

first: Myong-Hee

last: Sung

state: MD

address: 251 Bayview Boulevard, Room 06C226

department: National Institute on Aging

institute: NIH

data_processing:
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence using Cutadapt, mapped to reference human hg38 genome and gencode v30 transcripts, and NC_09333 (Human Herpes virus 8) using STAR software. Quantification was carried out with RSEM using transcriptome bam file created by star and gencode v24 transcriptome as index for RSEM.
Genome_build: hg38
Supplementary_files_format_and_content: RSEM

source_name: SLK cell line

growth_protocol: Cells are maintained in DMEM 10% FBS

extract_protocol:
Tissue was homogenized and total RNA was extracted by Direct-Zol miniprep kit.
Truseq Stranded RNA prep kit RS-122-2201

treatment_protocol: circRNA-expressing plasmids are transfected to SLK cell lines and selected with Hygromycin (0.5mg/ml) for 4 weeks.

tag: disease

value: carcinoma

value: SLK

tag: transfection


title: MCF10A, WTp53 cDNA insert, DMSO, Replicate 2 [RNAseq_7_DMSO_R2]

description: mRNA (RNA-seq) from MCF10A cells

tag: cell line

tag: cell type


title: Punicalagin-Treated-biological rep3

source_name: THP-1 treated with Punicalagin

value: Punicalagin


title: Punicalagin-Treated-biological rep1

value: passage 6


title: DMSO-treated-biological rep3

source_name: THP-1 treated with DMSO

value: DMSO


title: Punicalagin-Treated-biological rep2

value: passage 7


title: SLK-kcirc55_3


title: 29DXU82

value: ALS

value: peripheral blood

value: 2MIU IL2


title: 44AZD87


title: 98DAD98

value: Placebo


title: DMSO-treated-biological rep2


title: 25SJK94


title: 24ZOV85


title: SLK-kcirc55_1


title: 96OKC39


title: DMSO-treated-biological rep1


title: SLK-kcirc55_2


title: MCF10A, ∆40p53:WTp53 cDNA insert, 10uM Nutlin3a, Replicate 1 [41_Nutlin_p53_R1]

source_name: MCF10A, ∆40p53:WTp53 cDNA insert,10uM Nutlin3a

value: mammary gland; breast

value: epithelial

tag: crispr/cas9 insertion

tag: crispr/cas9 insertion location

value: 10uM Nutlin3a


title: MCF10A, ∆40p53:WTp53 cDNA insert, 10uM Nutlin3a, Replicate 2 [41_Nutlin_p53_R2]

tag: tissue

tag: treatment


title: 87FSD36


title: SLK-kcirc54_4


title: 85ORI83


title: CR2Crry-treated 28-day TBI brain 3

city: Charleston

first: Amer

last: Toutonji

state: SC

address: 173 Ashley Ave, CRI204

institute: Medical University of South Carolina

hyb_protocol: followed the manufacturer's instructions: NanoString (Seattle, WA)

data_processing: number of reads was normalized by housekeeping genes in the panel using nCounter® nSolver software

label: Nanostring

source_name: TBI-injured brain

label_protocol: the extract wasn't labeled

extract_protocol: mRNA was extracted from up to 1 mg of brain tissue using the RNeasy Lipid Tissue Mini Kit (Qiagen)

tag: injury

value: TBI (CCI)

tag: time point post tbi

value: day 28

value: CR2-Crry (16 mg/kg)

tag: age (weeks)


title: CR2Crry-treated 28-day TBI brain 1

scan_protocol: followed the manufacturer's instructions: NanoString (Seattle, WA)


title: CR2Crry-treated 28-day TBI brain 2


title: MCF10A, ∆40p53:WTp53 cDNA insert, 10uM Nutlin3a, Replicate 3 [41_Nutlin_input_R3]

description:
input DNA
processed data file: 41_Nutlin_0.0001_peaks_clean.narrowPeak

value: 3924bp:∆40p53 cDNA tethjered to full length p53 cDNA followed IRES expressed mCherry followed by SV40 polyA sequnece


title: CR2Crry-treated 7-day TBI brain 2

value: day 7


title: MCF10A, ∆40p53:WTp53 cDNA insert, 10uM Nutlin3a, Replicate 2 [41_Nutlin_input_R2]


title: CR2Crry-treated 7-day TBI brain 3


title: SLK-kcirc54_3

value: carcinoma

value: SLK


title: MCF10A, ∆40p53:WTp53 cDNA insert, 10uM Nutlin3a, Replicate 1 [41_Nutlin_input_R1]


title: 77WZO64


title: CR2Crry-treated 7-day TBI brain 1


title: 76UCI86


title: MCF10A, ∆40p53:WTp53 cDNA insert, DMSO, Replicate 3 [41_DMSO_p53_R3]

description:
DNA bound by p53
processed data file: 41_DMSO_0.0001_peaks_clean.narrowPeak

value: DO-1 (BD Biosciences, BD554293)


title: SLK-kcirc54_2


title: SLK-kcirc54_1


title: SLK-circGFP_4

value: circGFP


title: SLK-circGFP_3


description: Gene expression data from Normal urothelial tissue, patient B_AAN_04

source_name: Normal urothelial tissue from tumor surgery

tag: individual

value: patient B_AAN_04

tag: gender

value: female

tag: age at surgery

value: 74 years

value: normal urothelium


title: CR2Crry-treated 3-day TBI brain 3

value: day 3


title: CR2Crry-treated 3-day TBI brain 2


title: 76ABW87

tag: diagnosis

value: ALS

value: peripheral blood

value: Placebo

tag: time point


title: 57TOS36


title: MCF10A, ∆40p53:WTp53 cDNA insert, DMSO, Replicate 1 [41_DMSO_p53_R1]

value: mammary gland; breast

value: epithelial

tag: chip antibody


title: SLK-circGFP_1

tag: disease


title: MCF10A, ∆40p53:WTp53 cDNA insert, DMSO, Replicate 2 [41_DMSO_p53_R2]


title: HeLa -TRIzol

city: Beijing

first: Tong

last: Liu

address: Life Science Park Road

institute: State Key Laboratory of Proteomics

data_processing:
For the estimation of the rRNA content of libraries, reads were aligned to a collection of human ribosomal sequences which retrieved from NCBI nucleotide database. All reads were aligned to those sequences using bowtie2 and reads that failed to align were saved as non-rRNA reads file.
Reads in this file were aligned to the complete hg38 assembly by using STAR.Percentages of the rRNA content were estimated by comparing the number of reads aligning to rRNA sequences and residual reads aligning to the complete hg38 assembly.
For estimating the content of RNA biotypes in libraries, reads were aligned to the hg38 assembly using STAR.
counting was performed with HTSeq-count using the geneset annotated by GENCODE Release 21 (GRCh38) and using the GTF feature ‘gene’ for counting.
Genome_build: GRCh38
Supplementary_files_format_and_content: tab-delimited text file include raw reads count

source_name: HeLa cells

extract_protocol:
The RP-complexes from HeLa cells were captured by PP, proteinase K  was added to digest protein, the eluted RNAs were further purified by TRIzol following the manufacturer’s instructions. For other samples, RNA was harvested using Trizol reagent
RNA was used to construct the cDNA library using the TruSeq RNA Library Prep Kit v2 (Illumina, not stranded)

value: HeLa

tag: protocol

value: TRIzol


title: CR2Crry-treated 3-day TBI brain 1

value: TBI (CCI)

value: CR2-Crry (16 mg/kg)


title: SLK-circGFP_2

value: carcinoma

tag: cell line

value: SLK

tag: transfection


title: MCF10A, ∆40p53:WTp53 cDNA insert, DMSO, Replicate 3 [41_DMSO_input_R3]

description:
input DNA
processed data file: 41_DMSO_0.0001_peaks_clean.narrowPeak

tag: cell type

tag: crispr/cas9 insertion

value: 3924bp:∆40p53 cDNA tethjered to full length p53 cDNA followed IRES expressed mCherry followed by SV40 polyA sequnece


title: MCF10A, ∆40p53:WTp53 cDNA insert, DMSO, Replicate 2 [41_DMSO_input_R2]

tag: tissue

tag: crispr/cas9 insertion location


title: HeLa -PP enrichment

value: PP enrichment


title: MCF10A, ∆40p53:WTp53 cDNA insert, DMSO, Replicate 1 [41_DMSO_input_R1]

tag: treatment


title: 48BFT76


title: 22046_3b: SW480 cells YH-620 treatment rep2

city: Los Angeles

first: Kahn

last: Michael

address: NRT 4501, 1450 Biggy Street

department: Department of Biochemistry and Molecular Biology

institute: University of Southern California

data_processing:
Illumina bcl2fastq v2.18.0.12 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to hg38 whole genome using Tophat2 2.0.14 with default parameters
The HTSeq software (v.0.6.1) was applied to generate the count matrix, with default parameters. Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated using DEseq2 1.14.1
Genome_build: hg38
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample in RPKM

source_name: SW480 cell line

growth_protocol: SW480 cells were grown in DMEM (High Glucose, 10% FBS, Pen/Strep) in 5% CO2, 37 °C incubator.

extract_protocol: SW480 cells were removed from incubator and 1% oxygen chamber, and medium was aspirated, after which RNA was harvested using Trizol reagent.

treatment_protocol: SW480 cells were split/plated and incubated overnight at 37 °C, 5% CO2. Next, cells were transferred to a 1% oxygen chamber, and 1% oxygen gas was flowed through for 10 minutes, after which the chamber was clamp sealed and put into 5% CO2, 37 °C incubator and incubated for 24 hours. Next, spent medium was removed, and cells were treated with Control (DMSO), 10 uM YH-620, or 10 uM ICG-001 and then transferred back to the 1% oxygen chamber, and 1% oxygen gas was flowed through for 10 minutes, after which the chamber was clamp sealed and put into 5% CO2, 37 °C incubator and incubated for an additional 24 hours.

value: Colon

tag: disease state

value: Colorectal adenocarcinoma

value: Treated with YH-620


title: MCF10A, WTp53:WTp53 cDNA insert, 10uM Nutlin3a, Replicate 3 [21_Nutlin_p53_R3]

description:
DNA bound by p53
processed data file: 21_Nutlin_0.0001_peaks_clean.narrowPeak

value: 4041bp:full length p53 tethered to full length p53 cDNA followed IRES expressed mCherry followed by SV40 polyA sequnece

value: 10uM Nutlin3a

value: DO-1 (BD Biosciences, BD554293)


title: PBS-treated 28-day TBI brain 3

tag: injury

tag: time point post tbi

value: day 28

value: vehicle (PBS)

tag: age (weeks)


title: 34CJU23

value: ALS

value: peripheral blood

value: Placebo


title: PBS-treated 7-day TBI brain 3

value: TBI (CCI)

value: day 7


title: PBS-treated 7-day TBI brain 2


title: 22045_3a: SW480 cells YH-620 treatment rep1


title: PBS-treated 28-day TBI brain 2


title: MCF10A, WTp53:WTp53 cDNA insert, 10uM Nutlin3a, Replicate 2 [21_Nutlin_p53_R2]

source_name: MCF10A, WTp53:WTp53 cDNA insert,10uM Nutlin3a

value: mammary gland; breast

value: epithelial


title: MCF10A, WTp53:WTp53 cDNA insert, 10uM Nutlin3a, Replicate 1 [21_Nutlin_p53_R1]


title: 22044_2b: SW480 cells ICG-001 treatment rep2

value: Treated with ICG-001


title: PBS-treated 28-day TBI brain 1


title: 22043_2a: SW480 cells ICG-001 treatment rep1


title: PBS-treated 7-day TBI brain 1


title: PBS-treated 3-day TBI brain 2

value: day 3


title: MCF10A, WTp53:WTp53 cDNA insert, 10uM Nutlin3a, Replicate 3 [21_Nutlin_input_R3]

description:
input DNA
processed data file: 21_Nutlin_0.0001_peaks_clean.narrowPeak


title: MCF10A, WTp53:WTp53 cDNA insert, 10uM Nutlin3a, Replicate 2 [21_Nutlin_input_R2]


title: 22042_1b: SW480 cells Control treatment rep2

value: Colon

value: Treated with Control (DMSO)


title: 33UVN85


title: 95HNE98

value: 2MIU IL2


title: 93GXQ33


title: 82AII97


title: 33IEV23


title: 98HXI86


title: 56JIT63


title: 55WDE55


title: 46DHR29


title: PBS-treated 3-day TBI brain 1


title: MCF10A, WTp53:WTp53 cDNA insert, 10uM Nutlin3a, Replicate 1 [21_Nutlin_input_R1]

value: 4041bp:full length p53 tethered to full length p53 cDNA followed IRES expressed mCherry followed by SV40 polyA sequnece


title: MCF10A, WTp53:WTp53 cDNA insert, DMSO, Replicate 2 [21_DMSO_p53_R2]

description:
DNA bound by p53
processed data file: 21_DMSO_0.0001_peaks_clean.narrowPeak

source_name: MCF10A, WTp53:EWTp53 cDNA insert, DMSO


title: 38QCW65


title: 22041_1a: SW480 cells Control treatment rep1

value: Colorectal adenocarcinoma


title: mDA-BAG-2

city: BALTIMORE

first: Minseong

last: Kim

address: 733 NORTH BROADWAY, SUITE 763, Euodwood Apt 308

department: ICE

institute: JHMI

description: total RNA

data_processing:
Trimmomatic program is used to remove adapter sequences and bases with base quality lower than three from the ends. Also using sliding window method, bases of reads that does not qualify for window size 4, and mean quality 15 are trimmed. Afterwards, reads with length shorter than 36bp are dropped to produce trimmed data.
In order to map cDNA fragments obtained from RNA sequencing, UCSC hg19 was used as a reference genome. Table 3 shows the statistic obtained from HISAT2, which is known to handle spliced read mapping through Bowtie2 aligner. You can check number of processed reads, mapped reads.
Known genes and transcripts are assembled with StringTie based on reference genome model. After assembly, the abundance of gene/transcript is calculated in the read count and normalized value as FPKM (Fragments Per Kilobase of transcript per Million mapped reads) for a sample.
In order to reduce systematic bias, estimates the size factors from the count data and applies Relative Log Expression (RLE) normalization with DESeq2 R library. In case of DESeq2, read count+1 & Logarithm value is uesd to visualize the plots before normalization and regularized log (rlog) transformed value is used to visualize the plots after normalization.
Genome_build: hg19
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample
Supplementary_files_format_and_content: tab-delimited text files include raw count values for each Sample
Supplementary_files_format_and_content: tab-delimited text files include fold change values for each Sample

source_name: PD-iPSC derived mDA neuron

growth_protocol: We cultured undifferentiated SNCA triplication PD hiPSCs (ND27760-8) and opto-α-syn (AAVS1::opto-α-syn) PD hiPSCs on mitotically inactivated mouse embryonic fibroblasts (MEFs, Global Stem or Applied Stem Cell), in hESC medium containing DMEM/F-12, 20% knockout serum replacement (KSR), 0.1 mM MEM-NEAA, 2 mM L-glutamine, 55 μM β-mercaptoethanol (all from Life Technologies) and 10 ng/mL FGF2 (R&D Systems) as used routinely for iPSC cultures. All cells were maintained at 37 °C and 5% CO2 in a humidified incubator.

extract_protocol:
RNA was harvested using Trizol reagent. Illumina TruSeq Stranded mRNA Library Prep (Cat#20020594) was used with 1 ug of total RNA for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: For mDA neuron differentiation, Briefly, dissociated hiPSCs were plated on Geltrex at a density of 50,000 cells/cm2 in MEF-conditioned KSR medium containing DMEM/F-12, 20% KSR, 0.1 mM MEM-NEAA, 2 mM L-glutamine, and 55 μM β-mercaptoethanol with 10 ng/mL FGF2 and 10 μM ROCK-inhibitor (Y-27632, Cayman Chemical). After confluency of the cells reached 80%–90%, differentiation was initiated by switching to KSR medium supplemented with 100 nM LDN193189 (STEMCELL Technologies) and 10 μM SB431542 (Cayman Chemical). Supplements of 100 ng/mL Shh (C25II, R&D), 2 μM Purmorphamine (PMP, Cayman Chemical) and 100 ng/mL FGF8 (PeproTech) were added on days from 1 to 7, and 3 mM CHIR99021 (CHIR, Tocris) was added at day 3 to day 11. Beginning at day 5, the KSR medium was gradually replaced with increasing amounts of N2 medium(Oh et al., 2016) (25% increments every other day). To expand neural progenitors, the cells were split on Geltrex and maintained in medium containing DMEM/F-12, N-2 supplement (Life Technologies), 2 mM L-glutamine, 1% penicillin/streptomycin, 100 nM LDN193189, 3 µM CHIR and 10 µM Y-27632 on day 11. After that, the cells were re-plated on dishes pre-coated with Geltrex in NB/B-27 medium supplemented with 3 µM CHIR, 20 ng/mL BDNF, 0.2 mM ascorbic acid, 20 ng/mL GDNF, 1 ng/mL TGFβ3, 0.5 mM dbcAMP and 10 µM DAPT for at least 10 days to complete differentiation.

value: iPSC-derived mDA neurons

value: Bluelight + BAG 1uM for 24hrs


title: MCF10A, WTp53:WTp53 cDNA insert, DMSO, Replicate 3 [21_DMSO_p53_R3]

value: DO-1 (BD Biosciences, BD554293)


title: MCF10A, WTp53:WTp53 cDNA insert, DMSO, Replicate 1 [21_DMSO_p53_R1]


title: mDA-CDC-2

description: total RNA

value: Bluelight + CDC 1uM for 24hrs


title: Sham brain 2

source_name: normal mouse brain

value: sham

value: not applicable

value: vehicle (PBS)


title: mDA-BAG-1

description: total RNA


title: Sham brain 1