title: 32

data_processing:
The quality of the sequencing was verified with FastQC.
All the reads were trimmed using Cutadapt to remove the adaptor sequences.
The reads that have been trimmed were mapped against hg38 reference genome using STAR.
A table of read counts for quantifiable genes were generated using RSEM.
Differential gene expression analysis between groups of samples was performed using edgeR with batch effect correction due to inter-donor variability.
Genome_build: hg38
Supplementary_files_format_and_content: All processed data files are comma-separated values. Each row contains the Ensemble ID, the normalized counts in CPM and the corresponding gene name.

extract_protocol:
Total RNA was extracted by using Qiagen miRNeasy Micro Kit. Contaminating genomic DNA was removed by DNase digestion.
mRNA libraries were prepared using Lexogen QuantSeq 3' mRNA-Seq Library Prep Kit FWD.

treatment_protocol: HSPCs transduced with IRF8-ERT2-encoding construct were treated with compound solvent or 50 nM of 4-hydroxytamoxifen at day 0 (seeding) and day 3 post co-culture.

value: 3rd donor

value: compound solvent control (0 nM)


title: CD7m_IRF8ERT2_B

status: Public on Dec 11 2022

type: SRA

city: Ghent

first: Tom

last: Taghon

state: East Flanders

address: Corneel Heymanslaan 10

department: Diagnostic Sciences (GE32)

country: Belgium

institute: Ghent University

library_source: genomic

data_processing:
The quality of the sequencing was verified with FastQC.
Sequencing reads were trimmed using NGMerge.
Alignment was done using Bowtie2 (v2.2.6) with parameter --very-sensitive using the human hg38 reference genome.
Duplicate reads were removed using SAMTools.
Peak calling was performed with MACS2 (v2.1.2) and the option --no-model.
Significant peaks at an adjusted p-value cut-off of 0.05 were combined into one matrix across all samples, with merging of peaks that show at least 50 percent overlap.
The function SummarizeOverlaps of the GenomicAlignments package in R was used for read counting using this matrix and the BAM files.
DESeq2 was used to detect significantly differential opened chromatin sites between samples, using a design containing replicate and condition.
An adjusted p-value of 0.05 was used to retain significant hits where Homer findMotifsGenome was used to perform motif enrichment analysis.
Genome_build: hg38
Supplementary_files_format_and_content: DESeq2 normalized counts
Supplementary_files_format_and_content: Raw counts

molecule: genomic DNA

organism: Homo sapiens

source_name: co-cultured HSPCs

growth_protocol: lin-CD123-CD34+GFP+ transduced HSPCs were sorted and co-cultured with OP9-DLL4 stromal cells to induce T cell differentiation.

extract_protocol:
Tagmented genomic DNA was extracted by using Qiagen MinElute PCR purification kit.
DNA libraries were constructed as described by Corces et al., Nat. Methods (2017).

treatment_protocol: HSPCs transduced with IRF8-ERT2-encoding construct were treated with 50 nM of 4-hydroxytamoxifen at day 0 (seeding) and day 3 post co-culture.

tag: tissue

value: cord blood

tag: donor

value: B

tag: condition

tag: treatment

value: 50 nM of 4-hydroxytamoxifen

tag: phenotype


title: 33

library_source: transcriptomic

molecule: total RNA

value: 50 nM 4-OHT


title: CD7m_IRF8ERT2_A

value: A


title: 31

description: Normalized counts for this sample were derived from two comparisons: empty vector vs solvent control and empty vector vs 4-OHT.

value: empty vector control

value: untreated


title: B-813 [arid1a ko Arid1a ChIP-seq]

city: Tainan

first: Tzu-Lei

last: Kuo

state: Taiwan

address: No. 367, Sheng Li Road

department: Institute of Cancer Research

country: Taiwan

institute: Taiwan NHRI

data_processing:
The sequences were processed and analyzed by GENEWIZ.Data analysis  Quality Control:  In order to remove technical sequences, including adapters, polymerase chain reaction (PCR) primers, or fragments thereof, and quality of bases lower than 20, pass filter data of fastq format were processed by Cutadapt[1]（version 1.9.1）to be high quality clean data.  Mapping:  Firstly, reference genome sequences and gene model annotation files of relative species were downloaded from genome website, such as UCSC, NCBI, ENSEMBL. Secondly, bowtie-build was used to index reference genome sequence. Finally, clean data were aligned to reference genome via software Bowtie2 (version 2.2.6).  Peaks analysis:  Use MACS[2](V2) to analyze peaks quality control, peaks calling and peaks annotation.
Different peak analysis Combine the peaks of two different experimental groups with bedtools (version, 2.25.0) software, and then readcounts are counted in the combined peaks area. Difference analysis was performed using edgeR (V3.4.6) of the Bioconductor software package.
Assembly: mm10

organism: Mus musculus

source_name: primary Islet cell

growth_protocol: Islets were collected and pooled into a Petri dish with RPMI-1640 (GIBCO) containing 11.1 mM glucose, 100 units/ml penicillin, 100 μg/ml streptomycin, 2 mM Glutamax, 50 μg/ml gentamycin, 10 μg/ml Fungison and 10% fetal calf serum (Invitrogen)

extract_protocol:
Cells were fixed with 1% formaldehyde for 10 minutes. After quenching and cell lysis, chromatin was sonicated using a 60 Sonic Dismembrator (Fisher Scientifi c).
Next generation sequencing library preparations were constructed following the manufacturer’s protocol. For each sample, ChIP product was used for library preparation. The ChIP product was treated with End Prep Enzyme Mix for end repairing, 5’ Phosphorylation and dA-tailing in one reaction, followed by ligation to adaptors with a “T” base overhang. Adaptor-ligated DNA was then recovered using Beads.

tag: cell line

value: primary Islet cell

tag: cell type

value: Islet cell

tag: genotype

value: arid1a ko

tag: chip antibody

value: Arid1a antibody (Abnova, MAB15809)


title: C-813 [con HADC6 ChIP-seq]

value: con

value: HADC6 antibody (Cell signaling,#7612)


title: A-813 [con Arid1a ChIP-seq]


title: 52

value: 2nd donor


title: A3

value: 1st donor


title: 51


title: A2


title: A1


title: MDA::ER::pCDNA_siFOXA2 ER ChIP

city: Rennes

first: Raphaël

last: Métivier

address: Campus de Beaulieu

country: France

institute: CNRS UMR 6290

description: Chipped

data_processing:
Basecalls were performed using CASAVA (v1.8. or v1.8.2) to produce fastq files. The multiple files generated were the catenated into a single one.
Reads were aligned to the hg19 reference genome assembly using bowtie (Langmead et al., Genome Biol., 2009) with parameters allowing at most one mismatch and selected for unique mapping onto the genome.
Extracted reads were  converted to .wig signal files using samtools (Li et al., Bioinformatics, 2009) and MACS (Zhang et al., Genome Biol, 2008) with default parameters, incluing a normalization step towards the input sequences.
To minimize the bias of diverging sequencing depths between different ChIP samples prepared with the same antibody, the signal intensities of a given .wig were linearly normalized so as to be comparable to the .wig file with the highest sequencing depth.
Peak callings were then performed using the algorithm described in Serandour et al. (Nucleic Acids Research, 2012) with a p-value for the threshold set to 1e-8 and peaks defined as being constituted of at least 4 adjacent signals within 65 bp above the threshold values
Genome_build: hg19
Supplementary_files_format_and_content: .wig files containing MACS (v1.4) signals normalized to input reads

source_name: breast cancer cell line

growth_protocol: MCF7, MDA-MB231  MDA::ER::pCDNA and MDA::ER::FOXA1 cells were maintained  in DMEM (Sigma)/10% FCS (BioWest) supplemented or not with selection antibiotics for integrated vectors (0.8 mg/ml Hygromycin, 5 µg/ml blasticidin, 75 µg/ml Zeocin ).

extract_protocol:
Cells were and lysed for 10 min in ice in 300 µl of lysis buffer [10 mM EDTA, 50 mM Tris-HCl (pH 8.0), 0.1% SDS, 0.5% Empigen BB (Sigma)]. Crosslinked chromatin was sheared to an average size of 400 bp by sonication for 15 min with 30secs on/off cycles at full power in a BioRuptor apparatus (Diagenode). Following 10 minutes of centrifugation at 10,000 x g, 30 µl were kept as input fraction and the remainder diluted 5 fold in IP buffer (2 mM EDTA, 100 mM NaCl, 20 mM Tris-HCl [pH 8.1], and 0.5%Triton X-100) before proceeding to immunoprecipitation overnight at 4°C under rocking using 2 µg of antibodies and 5 µg of yeast tRNAs. Complexes were recovered after 3 hr incubation at 4°C with 5 µg of yeast tRNAs and 100 µl of a 50% protein A or protein G-Sepharose beads slurry (Amersham Pharmacia Biosciences). Precipitates were then serially washed, using 500 µl of Washing Buffers (WB) I [2 mM EDTA, 20 mM Tris-HCl (pH 8.1), 0.1% SDS, 1% Triton X-100, 150 mM NaCl], WB II [2 mM EDTA, 20 mM Tris-HCl (pH 8.1), 0.1% SDS, 1% Triton X-100, 500 mM NaCl], WB III [1 mM EDTA, 10 mM Tris-HCl (pH 8.1), 1% NP-40, 1% Deoxycholate, 0.25 M LiCl] and then twice with 1 mM EDTA, 10 mM Tris-HCl (pH 8.1). Precipitated complexes were removed from the beads through two sequential incubations of 10 min with 50 µl of 1% SDS, 0.1 M NaHCO3. Crosslink was reversed by an overnight incubation at 65°C with 50 µg of Proteinase K (Sigma). Following a subsequent incubation of the samples with 2.5 µg RNAse (Sigma) for 1h at 37°C, the DNA was then purified on NucleoSpin™ columns (Macherey-Nagel) using NTB buffer and eluted in 40 µl of elution buffer. Efficiency of the ChIP assay was evaluated using qPCR positive and negative controls before proceeding to subsequent steps. For each ChIP conditions, from 10 to 18 individually prepared samples were pooled and submitted for librairy preparation.
Libraries were prepared according to Illumina's instructions by IGBMC sequencing facility (Strasbourg, France)

treatment_protocol: Following 3 days of culture in DMEM/0.5% charcoal-stripped serum (BioWest), 15.10E6 MDA::ER or MDA-MB231 cells were treated for 50 minutes with 10-8M final estradiol and fixed for 10 min at room temperature in a PBS solution containing 1.5% formaldehyde (ACROS). After a 5 min quenching with 0.125M glycine, cells were washed twice in ice-cold PBS and collected in 1 ml of cold buffer (100 mM Tris-HCl [pH 9.4] and 100 mM DTT). The procedure was identical in the case of MCF-7 cells, but these cells were first cultivated for 48h in DMEM/10% serum before the 3 days in stripped serum. Expression of the FOXA1 transgene in MDA::ER::FOXA1 cells was induced 48h before the E2 treatment by adding Tetracycline (Sigma) at a final concentration of 1.5 microg/ml. Transfection of siRNAs was performed on 2.5.10E6 MDA::ER::pCDNA cells  plated in 9cm dishes in DMEM/5%FCS for 16h and then grown for 24h in DMEM/2.5% csFCS. Media was then replaced with 4 ml of FCS free Opti-MEM (Sigma) and 1 microM siRNAs were then transfected using oligofectamine as recommended by the manufacturer (Invitrogen). Following 6h of incubation, the media was completed with 125 microl of csFCS, and E2 stimulation (10-8M) was done 36h later.

tag: gender

value: female

value: breast cancer

tag: integrated vector

tag: transfected sirna

value: yes:siRNA directed against FOXA2

tag: hormone treatment

value: 10-8M Estradiol (Sigma)

value: Antibody Mix:HC20 (Santa Cruz) + ab10 (TE111-SD11, NeoMarker)


title: MDA::ER::pCDNA_siLuc ER ChIP

description: Chipped

value: yes:control siRNA directed against Luciferase


title: MDA-MB231 H3K27ac ChIP

description: Chipped

value: no

value: ab4729 (Abcam)


title: 53

value: cord blood

value: 50 nM 4-OHT


title: MDA-MB231 H3K4me2 ChIP

description: Chipped

value: 07-030 (Millipore)


title: MCF-7 H3K27ac ChIP

description: Chipped


title: MDA::ER::FOXA1_EtOH_3

type: RNA

description: This sample is of MDA::ER::FOXA1 cells (MDA-MB231 cells engineered to constitutively express the estrogen receptor and to express FOXA1 in the presence of Tetracyclin) treated with ethanol (vehicle control) for 4h. Third of the three independent replicates used in this experiment, each constituted by a pool of three separate cultures.

hyb_protocol: Hybridization was performed by NimbleGen Systems (Services, Reykjavic), following their standard operating protocol. See www.nimblegen.com.

scan_protocol: Standard NimbleGen protocols (scanned at NimbleGen services, Reykjavic). See www.nimblegen.com.

data_processing: Raw data (.pair) were subjected to the RMA algorithm (Robust Multi-Array Analysis; Irizarry et al. Biostatistics 4(2):249) with quantile settings (Bolstad et al. Bioinformatics 19(2):185) for background correction and normalization, as implemented within the ArrayStar software package (DNAstar, Inc.).

source_name: MDA::ER::FOXA1 cells treated for 4h with 0.1% ethanol (vehicle control)

label_protocol: Labeling was performed by NimbleGen Systems (Services, Reykjavic), following their standard operating protocol. See www.nimblegen.com.

growth_protocol: Cells were maintained in DMEM (Sigma) supplemented with 5% fetal calf serum (FCS, BioWest) and antibiotics (Roche) at 37°C under 5% CO2. Media was  complemented with 0.8 mg/ml Hygromycin (Calbiochem), 5 µg/ml Blasticidin (Invitrogen) and  75 µg/ml Zeocin (Invitrogen) for continuous selection of pCDNA3.1-Hygro-ER, pcDNA6/TR and empty pcDNA4/TO or pcDNA4/FOXA1 vectors, respectively.

extract_protocol: Total RNA was extracted using the RNeasy Mini kit (Qiagen) with homogenization facilitated by QIAShredders (Qiagen). Integrity and purity of the RNAs were controlled on an Agilent Bioanalyzer using the RNA 6000 Nano Assay (Agilent). Ten micrograms of selected samples exhibiting a RIN >9.8 were then subjected to cDNA synthesis using the Superscript Double-Stranded cDNA Synthesis Kit (Invitrogen) and 50 pmol random hexamers. cDNAs were then treated for 10 min at 37°C with 5 μg RNaseA (Invitrogen), purified through a phenol:chloroform:isoamyl alcohol extraction on Phase Lock light gels (Fisher Scientific) and then precipitated.

treatment_protocol: Cells were plated at 2x106 cells per 9cmØ dishes in DMEM/2.5% FCS (complemented with 0.8 mg/ml hygromycin, 5 µg/ml Blasticidin and  75 µg/ml Zeocin) for 16h. Following two washes with PBS, cells were then cultivated for 3 days in DMEM/0.5% charcoal-stripped serum (BioWest) complemented with all antibiotics. Confluent cells were then treated with Estradiol (E2, Sigma) at a final concentration of 10-8 M for 4h or with equivalent amounts of ethanol vehicle (0.1%).

value: breast cancer cells

tag: er status

value: Positive

tag: integrated vectors

value: pCDNA3.1-Hygro-ER + pcDNA6/TR + pcDNA4/TO/FOXA1


title: MDA::ER::FOXA1_E2_3

description: This sample is of MDA::ER::FOXA1 cells (MDA-MB231 cells engineered to constitutively express the estrogen receptor and to express FOXA1 in the presence of Tetracyclin) treated with 10-8M E2 for 4h. Third of the three independent replicates used in this experiment, each constituted by a pool of three separate cultures.

source_name: MDA::ER::FOXA1 cells treated for 4h with 10-8M estradiol


title: MDA::ER::FOXA1_EtOH_1

description: This sample is of MDA::ER::FOXA1 cells (MDA-MB231 cells engineered to constitutively express the estrogen receptor and to express FOXA1 in the presence of Tetracyclin) treated with ethanol (vehicle control) for 4h. First of the three independent replicates used in this experiment, each constituted by a pool of three separate cultures.


title: MDA::ER::FOXA1_E2_2

description: This sample is of MDA::ER::FOXA1 cells (MDA-MB231 cells engineered to constitutively express the estrogen receptor and to express FOXA1 in the presence of Tetracyclin) treated with 10-8M E2 for 4h. Second of the three independent replicates used in this experiment, each constituted by a pool of three separate cultures.


title: MDA::ER::pcDNA_E2_2

description: This sample is of MDA::ER::pcDNA cells (MDA-MB231 cells engineered to express the estrogen receptor and expressing the Tet Repressor) treated with 10-8M E2 for 4h. Second of the three independent replicates used in this experiment, each constituted by a pool of three separate cultures.

source_name: MDA::ER::pcDNA cells treated for 4h with 10-8M estradiol

value: pCDNA3.1-Hygro-ER + pcDNA6/TR + empty pcDNA4/TO


title: MDA::ER::FOXA1_EtOH_2

description: This sample is of MDA::ER::FOXA1 cells (MDA-MB231 cells engineered to constitutively express the estrogen receptor and to express FOXA1 in the presence of Tetracyclin) treated with ethanol (vehicle control) for 4h. Second of the three independent replicates used in this experiment, each constituted by a pool of three separate cultures.


title: MDA::ER::FOXA1 H3K27ac ChIP

description: Chipped


title: MCF-7 ER ChIP

description: Chipped


title: MDA::ER::FOXA1 H3K4me2 ChIP

description: Chipped


title: MDA::ER::FOXA1 FOXA1 ChIP

description: Chipped

value: ab 5089 (Abcam)


title: MCF-7 FOXA1 ChIP

description: Chipped


title: MDA::ER::pcDNA_E2_1

description: This sample is of MDA::ER::pcDNA cells (MDA-MB231 cells engineered to express the estrogen receptor and expressing the Tet Repressor) treated with 10-8M E2 for 4h. First of the three independent replicates used in this experiment, each constituted by a pool of three separate cultures.


title: MDA::ER::FOXA1_E2_1

description: This sample is of MDA::ER::FOXA1 cells (MDA-MB231 cells engineered to constitutively express the estrogen receptor and to express FOXA1 in the presence of Tetracyclin) treated with 10-8M E2 for 4h. First of the three independent replicates used in this experiment, each constituted by a pool of three separate cultures.


title: MDA::ER::pcDNA_EtOH_1

description: This sample is of MDA::ER::pcDNA cells (MDA-MB231 cells engineered to express the estrogen receptor and expressing the Tet Repressor) treated with ethanol (vehicle control) for 4h. First of the three independent replicates used in this experiment, each constituted by a pool of three separate cultures.

source_name: MDA::ER::pcDNA cells treated for 4h with 0.1% ethanol (vehicle control)


title: MDA::ER::pcDNA_EtOH_2

description: This sample is of MDA::ER::pcDNA cells (MDA-MB231 cells engineered to express the estrogen receptor and expressing the Tet Repressor) treated with ethanol (vehicle control) for 4h. Second of the three independent replicates used in this experiment, each constituted by a pool of three separate cultures.


title: MDA::ER::pCDNA H3K27ac ChIP

description: Chipped


title: C646+CPTH2

city: Fukui

first: Masaya

last: Nagaya

address: 3-9-1 Bunkyo

country: Japan

institute: University of Fukui

hyb_protocol: Samples were hybridized using Affymetrix hybridization kit materials • Heat cocktails at 99° for 5 minutes, then 45° for 5 minutes centrifuge at max speed for 1 minute. • Transfer 130μl of hyb solution to each array, then tape holes and parafilm. • Hybridize 16 hours at 45° at 60rpm• Fluidics washing is FS450_0002

scan_protocol: Gene ChIP Scanner 3000 7G System

data_processing:
Data were processed using R. MAT background correction were applied to probe intensities, gene level expression were summarized from the corrected intensities and then quantile normalized.
RaGene-2_0-st.pgf
RaGene-2_0-st.mps

label: biotin

organism: Rattus norvegicus

source_name: Rat lense, 30mM galactose, 6days, 40µM C646 & 80µM CPTH2 added

label_protocol: Samples were enzymatically fragmented and biotinylated using the WT Terminal Labeling Kit (Affymetrix)

extract_protocol: The lens removed from the culture medium was ground with a biosmasher until it fell apart. Then treat the sample with TRIZOL, cDNA synthesis and amplification was performed

tag: strain

value: Sprague Dawley

value: lens


title: CBP30

source_name: Rat lense, 30mM galactose, 6days, 10µM CBP30 added


title: MDA::ER::pcDNA_E2_3

description: This sample is of MDA::ER::pcDNA cells (MDA-MB231 cells engineered to express the estrogen receptor and expressing the Tet Repressor) treated with 10-8M E2 for 4h. Third of the three independent replicates used in this experiment, each constituted by a pool of three separate cultures.

value: female


title: CPTH2

source_name: Rat lense, 30mM galactose, 6days, 80µM CPTH2 added


title: TH1834

source_name: Rat lense, 30mM galactose, 6days, 50µM TH1834 added


title: MDA::ER::pCDNA FOXA2 ChIP

description: Chipped

value: breast cancer

value: Antibody Mix:clone 3C10 (Millipore) + ab23630 (Abcam)


title: MDA::ER::pCDNA H3K4me2 ChIP

description: Chipped

value: 10-8M Estradiol (Sigma)


title: CBP30+CPTH2

source_name: Rat lense, 30mM galactose, 6days, 10µM CPTH2 & 80µM CPTH2 added


title: MDA::ER::pcDNA_EtOH_3

description: This sample is of MDA::ER::pcDNA cells (MDA-MB231 cells engineered to express the estrogen receptor and expressing the Tet Repressor) treated with ethanol (vehicle control) for 4h. Third of the three independent replicates used in this experiment, each constituted by a pool of three separate cultures.


title: Galactose day6-3

source_name: Rat lense, 30mM galactose, 6days, replicate3


title: MDA::ER::pCDNA ER ChIP

description: Chipped

value: Antibody Mix:HC20 (Santa Cruz) + ab10 (TE111-SD11, NeoMarker)


title: BMSBF3_D712_S12

city: Burlington

first: Julie

last: Dragon

state: VT

address: 95 Carrigan Dr

department: MMG

country: USA

institute: UNIVERSITY OF VERMONT & ST AGRIC COLLEGE

data_processing:
Illumina Casava1.7 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 STAR 2.6 aligner
Sequenced reads were quantified using an Expectation-Maximization model, and translated to genes
Genes that have fewer than 30 counts were then filtered, and remaining genes median ratio plus 1 normalized, and differential gene expression was evaluated using the algorithm in DESeq2 v3.5 base on median ratio normalization and the negative binomial distribution of gene signal
Genome_build: hg38
Supplementary_files_format_and_content: tab-delimited text files include raw counts for each sample based on the Expectation Maximization algorithm (EM)

source_name: Myeloma Cell Line

growth_protocol: MM.1S cells were grown in RPMI-1640 media containing FBS and antibiotic/antimycotic

extract_protocol:
Total RNA was harvested in QIAZOL and prepared via Qiagen miRNEASY Kit with DNase On-column digestion (Qiagen; Hildnen, Germany)
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: MM.1S cells were cultured for 24 hours with 50 μM BMS309403, 50 μM SBFI-26 or the combination in a T25 at a density of 2.5x10^6 cells.

value: Myeloma Cell Line

tag: cell subtype

value: Malignant plasma cell


title: MCF-7 input

description: Input

value: no


title: BMSBF2_D711_S11


title: Galactose day6-1

source_name: Rat lense, 30mM galactose, 6days, replicate1

tag: tissue


title: C646

source_name: Rat lense, 30mM galactose, 6days, 40µM C646 added


title: control (w1118) heads at ZT16, sample 1 [ZT16WT1]

city: Lausanne

first: Tilmann

last: Achsel

state: VD

address: Rue Bugnon 9

department: Dept. Fundamental Neurosciences

country: Switzerland

institute: University of Lausanne

description:
htseq.genes.rpkm.txt
Supplementary Data 1.xlsx

data_processing:
Purity-filtered reads were adapters and quality trimmed with Cutadapt (v. 1.8, Martin 2011).
Reads matching to ribosomal RNA sequences were removed with fastq_screen (v. 0.9.3).
Remaining reads were further filtered for low complexity with reaper (v. 15-065, Davis et al. 2013).
Reads were aligned against Drosophila melanogaster.BDGP6.86 genome using STAR (v. 2.5.2b, Dobin et al. 2013).
The number of read counts per gene locus was summarized with htseq-count (v. 0.6.1, Anders et al. 2014) using Drosophila melanogaster.BDGP6.86 gene annotation.
Assembly: BDGP6.86
Supplementary files format and content: tab-delimited text files include RPKM values for each Sample
Supplementary files format and content: Excel file with statistic evaluation as published in the accompanying paper.

organism: Drosophila melanogaster

source_name: head

growth_protocol: Flies were maintained on standard cornmeal fly food at 25 °C, 60–80% relative humidity, in a 12-hrs light/dark cycle.

extract_protocol:
Fly heads were quickly collected at the specific time points using liquid nitrogen. Total RNA was isolated using Trizol Reagent (Gibco) according to the manufacturer’s instructions to extract total RNA.
RNA-seq libraries were prepared using 500 ng of total RNA and the Illumina TruSeq Stranded mRNA reagents (Illumina; San Diego, California, USA) on a Sciclone liquid handling robot (PerkinElmer; Waltham, Massachusetts, USA) using a PerkinElmer-developed automated script.

treatment_protocol: Flies were taken at ZT16

value: head

value: w1118 wild-type


title: control (w1118) heads at ZT16, sample 3 [ZT16WT3]


title: control (w1118) heads at ZT16, sample 2 [ZT16WT2]


title: MDA::ER::FOXA1 ER ChIP

description: Chipped


title: Galactose day6-2

source_name: Rat lense, 30mM galactose, 6days, replicate2


title: MDA::ER input

description: Input


title: BMSBF1_D710_S10


title: SBF3_D709_S9


title: Cyfip mutant heads at ZT16, sample 3 [ZT16HET3]


title: SBF2_D708_S8


title: Galactose day4-2

source_name: Rat lense, 30mM galactose, 4days, replicate2


title: Galactose day4-3

source_name: Rat lense, 30mM galactose, 4days, replicate3


title: Galactose day4-1

source_name: Rat lense, 30mM galactose, 4days, replicate1


title: Control

source_name: Rat lense, without galactose, 4 days


title: SBF1_D707_S7


title: Cyfip mutant heads at ZT16, sample 2 [ZT16HET2]


title: Cyfip mutant heads at ZT16, sample 1 [ZT16HET1]


title: p055_subassembly_oBC_devCRE

city: Montreal

first: Jean-Benoit

last: Lalanne

state: QC

address: 5155 Chem. de la Rampe

department: Biochemistry and Molecular Medicine

country: Canada

institute: University of Montreal

description:
Plasmid p055 oBC-CRE subassembly
Escherichia coli (for plasmid amplification)
p055_oBC_CRE_subassembly.txt
final_subassembly_oBC_devCRE_mBC_triplets_mEB_series.txt

data_processing:
p025 promoter series:
Sequencing data was demultiplexed using bcl2fastq. Raw fastq files were processed first by trimming unnecessary cycles from the 3’ end (10 cycles from read 1, 5 cycles from read 2, 9 cycles from index 1) using seqtk. Forward and reverse oBC reads were joined and error corrected with PEAR (options -v 16 -m 16 -n 16 -t 16). Using custom python and R scripts, assembled oBC reads were combined with mBC reads, and oBC/mBC pairs were counted. The read count distribution displayed a clear bimodal distribution, and oBC-mBC pairs with >500 reads were retained as valid. In order to further restrict the list of oBC-mBC pairs unique across the five bottlenecked libraries, all oBC/mBC pairs were combined, and any pair constituted from a oBC or mBC appearing more than once (either within a library, or across different libraries) was discarded to avoid mapping conflicts in the analysis of single-cell reporter data (amounting to 24% of high read count pairs), leaving 1122 unique oBC-mBC pairs across the five libraries (number of oBC-mBC pairs per library ranging from 139 to 306, with a median of 205).
p025 recloned complex:
Sequencing data was demultiplexed using bcl2fastq. Raw fastq files were processed first by trimming unnecessary cycles from the 3’ end using seqtk (https://github.com/lh3/seqtk). Forward and reverse oBC reads were joined and error corrected with PEAR81 (options -v 16 -m 16 -n 16 -t 16). Using custom python and R scripts, assembled oBC reads were combined with mBC reads, and oBC/mBC pairs were counted. The read count distribution displayed a clear bimodal distribution, and oBC-mBC pairs with >5 reads were retained as present in the library (~1.2M oBC-mBC pairs). To mark possible non-uniquely paired barcodes, we computed the proportion of read counts to each oBC and mBC from a given oBC-mBC pair. oBC-mBC pairs with oBC or mBC whose read counts proportion belonging to the pair of less than 95% were marked as likely non-unique (78.1% likely unique pairs by this criterion).
p055 oBC-CRE:
Following demultiplexing (from the P7 index), the sequencing data was processed by first aligning read 1 (mapping to CRE) using bowtie2 (v2.4.4) using option ‘-k 2’ to reporter multi-mapping regions (some of our devCRE segments overlapped given the proximity of the called peaks and extension from 500 bp to ≈1 kb tested regions). The resulting alignment sam file was then sorted, converted to bam using SAMtools, and merged with the oBC (read 2) using custom scripts into a file storing the oBC, CRE identity of the mapping, position and strand of aligned read within the CRE. Read counts to each oBC-CRE pair were then piled up with custom scripts, retaining information about distribution of alignment positions and strand within the CRE for downstream processing.
The piled-up count data on oBC-CRE pairs was then filtered to identify bona fide, unique pairs. First, pairs with median mapping position outside the expected range from the size exclusion step (<30 bp and >300 bp) and mapping on the incorrect strand were filtered out. Then, the proportion of oBC reads mapping to any given CRE was calculated across oBC-CRE pairs, and only pairs with >95% of oBC reads mapping to a unique CRE were retained. The read count distribution across all oBC-CRE pairs was bimodal, and only pairs with >30 reads (between the two modes) were retained. Finally, pairs with anomalously small or large mapping position dispersal (90th to 10th percentile difference mapping positional spread <30 bp or >300 bp) were filtered out. We note that the positional filters enabled unambiguous discrimination between different overlap CREs (given that in all cases one of the CRE would have out of range mapping positions compared to the expected size from the amplicon library). Two elements (Gata4:chr14_5749, Txndc12:chr4_7975) shared a short identical sequence complicating the mapping, and were treated separately to not confound the fraction of oBC reads mapping to a given CRE. Following these filtering steps, we were left with 43.6k valid oBC-CRE pairs.
These oBC-CRE subassembled pairs were then linked with the previously determined oBC-mBC pairs from the starting plasmid library p025.  Briefly, oBC (from final oBC-CRE pairs) were joined to mBC via valid oBC-mBC pairs (restricting to the uniquely mapped pairs). The resulting valid triplets oBC-CRE-mBC were then joined with the oBC-promoter-mBC triplets of the exogenous promoter library, and any oBC or mBC appearing  in both libraries were removed from the final triplet list as ambiguous. The final number of valid oBC-CRE-mBC triplets was 33.0k, with a median of 145 valid mBC-oBC pairs per CRE.
Assembly: N/A
Supplementary files format and content: p025_recloned_complex_mBC_oBC_subassembly: Read count table for the different oBC-mBC pairs identified in the recloned p025 complex library. Column 1: oBC; column 2: mBC; column 3: oBC-mBC pair read counts; column 4:  proportion of read counts; column 5: sum of read counts with that oBC; column 6: sum of read counts with that mBC; column 7: proportion of reads with that oBC from that specific oBC-mBC pair; column 8: proportion of reads with that mBC from that specific oBC-mBC pair; column 9: boolean if column 7 > 0.95 (oBC likely uniquely represented);  column 10: boolean if column 8 > 0.95 (mBC likely uniquely represented); column 11: boolean if both oBC and mBC uniquely represented.
Supplementary files format and content: p055_oBC_CRE_subassembly: Count table for the valid assembled oBC-CRE pairs from plasmid library p055. Column 1: reverse complement oBC; column 2: CRE identity; column 3: number different tagmented positions (serving as a pseudo UMI); column 4: total read count for this oBC-CRE pair; column 5: total number of reads matched to specific oBC; column 6: proportion of reads associated with this specific oBC coming from the oBC-CRE pair (only oBC-CRE pairs with this fraction above 0.95 are retained as uniquely paired).
Supplementary files format and content: final_subassembly_oBC_promoters_mBC_triplets_promoter_series: The processed file contains the final list of 1122 oBC-mBC pairs passing filters (column 1: class of CREs, here all promoters; column 2: identity of CRE [here one of the five promtoers]; column 3: oBC; column 4: RC_oBC; column 5: mBC; column 6: RC_mBC).
Supplementary files format and content: final_subassembly_oBC_devCRE_mBC_triplets_mEB_series: contains the final valid oBC-CRE-mBC triplet both for exogenous promoters and putative devCRE tested. column 1: CRE_class (promoters: exogenous promoters, devCRE: putative devCRE); column 2: CRE identity; column 3: oBC; column 4: reverse complement oBC; column 5: mBC; column 6: reverse complement mBC.

organism: Escherichia coli

source_name: bacterial cell

extract_protocol:
Plasmids were purified with Zymo MidiPrep kit.
p025 promoter series: small pools were generated by separately bottlenecking a ~12k clones library to an estimated 300 clones five separate times, and the oBC and mBC were subassembled for the separate pools. Briefly, amplicons were generated from the bottlenecked p025 as template, and using primers o345 and o337-o341 (indexed primer, one per library). Reactions were carried out in 50 uL volume with 20 ng input plasmid template (25 uL polymerase master mix, 2.5 uL 10 uM o345, 2.5 uL 10 uM indexed primer o337-o341, 0.25 uL 100x SYBr green, water to 50 uL) using Kapa HiFi PCR master mix (Roche) with PCR conditions: 95C 3 min, cycling with 98C 20 seconds, 60C 20 seconds, 72C 30 seconds. Reaction was tracked by qPCR and collected at the inflection point. Amplicons were purified by 1x Ampure.
Complex recloned p025: Amplicons were generated as above from p025 as template, and using primers o345 and o337. Reactions were carried out in 50 uL volume with 20 ng input plasmid template (25 uL polymerase master mix, 2.5 uL 10 uM o345, 2.5 uL 10 uM indexed primer o337, 0.25 uL 100x SYBr green, water to 50 uL) using Kapa HiFi PCR master mix (Roche) with PCR conditions: 95C 3 min, cycling with 98C 20 seconds, 60C 20 seconds, 72C 30 seconds. Reaction was tracked by qPCR and collected at the inflection point. Amplicons were purified by 1x Ampure.
Plasmid library p055 was tagmented with Tn5 at a concentration such that the expected fragment size would be larger than the oBC to minP distance (≈1.3 kb), determined by a Tn5 titration curve experiment. Following tagmentation (5 uL 2x TD buffer, 0.4 uL Tn5 NexteraV2 enzyme 1, 3.6 uL water, 1 uL 10 ng/uL plasmid library; 30 min at 37C), the fragmented plasmids were cleaned up (Zymo clean and concentrator, 3:1 binding buffer), eluted in 10 uL Tris 8 10 mM. 1 ng (1 uL of the elution) was amplified via semi-specific PCR with a Nextera primer with a P5 handle (o512, binding to all P5 tagmentation events) and a oBC upstream primer (o502, binding to specific portion of the plasmid) in 25 uL (8.9 uL water, 12.5 uL 2x NEBNext master mix, 1.25 uL 10 uM o502, 1.25 uL o512, 1 uL tagmented plasmids, 0.1 uL 200x SYBR green) with the following conditions (gap fill: 72C for 5 min, 98C for 30 sec, then 12 cycles of 98C for 10 sec, 65C for 30 sec, 72C for 1 min). Following purification (Zymo clean and concentrators), the amplified libraries were run on PAGE (6% TBE, 180V, 30 min). Semi-specific o502+o512 products between 450 bp and 800 bp were size selected on the gel, purified (minimum size size from CRE ≈ 75 bp), and sequenced.
Read structure (p025 promoter series): read 1 o346 (oBC, 26 cycles), index 1 o347 (library index, 6 cycles), read 2 o348 (oBC reverse complement, 25 cycles), index 2 o349 (mBC reverse complement, 20 cycles)
Read structure (p025 recloned complex): read 1 o346 (oBC, 30 cycles); index 1 o347 (library index, 15 cycles); read 2 o334 (mBC reverse complement, 18 cycles); index 2 o348 (oBC reverse complement)
Read structure (p055: read 1: CRE sequence, illumina Nextera primer [not custom], 34 cycles; index 1: P7-idx, primer o432 15 cycles; read 2: oBC, primer o433, 30 cycles
Custom amplicons for reporter constructs subassemblies

value: bacterial cell


title: BMS1_D704_S4


title: MM1_D701_S1


title: MM2_D702_S2


title: siWTAP+H. pylori.Input

city: Hong Kong

first: Judeng

last: ZENG

address: Room 902, Li Ka Shing Medical Science Building, Prince of Wales Hospital

country: China

institute: The Chinese University of Hong Kong

description:
Cells transfected with 20 nM WTAP siRNAs were co-cultured with H. pylori (MOI 1:100) for 24 hours.
m6A-enriched RNA

data_processing:
The reads from mRNA input and m6A IP sequencing libraries were aligned to hg19 reference genome using Tophat.
Both MACS/MACS2 and exomePeak were used to call m6A peaks based on the bam files.
To achieve high specificity, only the m6A peaks called by both MACS / MACS2 and exomePeak were retained for further analysis.
Differentially methylated m6A peaks were identified according to the procedure described by Schwartz et al. (PMID: 24269006).
Sequence motifs enriched in m6A peak regions compared to control regions were identified using DREME.
Assembly: hg19
Supplementary files format and content: Column A ~ E: the coordinates of the methylated RNA sites in bed format, please ref http://genome.ucsc.edu/FAQ/FAQformat.html#format1;   Column F: the length of the methylated RNA sites;   Column G ~ H: the transcript id and gene symbol of methylated RNA;   Column I ~ K: the enrichment results calculated by MACS;   Column L ~ S: the annotation of the methylated gene.
Supplementary files format and content: Legend:   transcript_id: the transcript id;   GeneName: the symbol of the methylated gene;   -10*log10(pvalue): -10*log10(pvalue) for the peak;   fold_enrichment: fold enrichment for the peak;   FDR: False discovery rate;   GeneID: the unique identifier for a gene in NCBI's database;   Synonyms: the default symbol for the gene;   dbXrefs: bar-delimited set of identifiers in other databases for this gene;   description: a descriptive name for this gene;   Pathway: the kegg pathways that include the gene;   GO_CC: the gene ontologies of cellular component category that include the gene;   GO_MF: the gene ontologies of molecular function category that include the gene;   GO_BP: the gene ontologies of biological process category that include the gene.
Library strategy: m6A-seq

growth_protocol: The stored (at -80℃) H. pylori strains were spread on the horse blood agar plates (Thermo Fisher, PB0122A) and grown in an anaerobic jar with a microaerobic environment (Oxoid CampyGen 2.5L Sachet, Thermo Fisher, CN0025A) at 37℃ for 4 - 5 days. Human gastric epithelial cell line HFE145 was cultured in Dulbecco’s Modified Eagle Medium (DMEM, Gibco, Thermo Fisher, 11995065) supplemented with 10% Fetal Bovine Serum (FBS, Gibco, Thermo Fisher, A4766801) at 37℃ with 5% CO2.

extract_protocol:
Total RNA was extracted form treated cells using RNAiso reagent (Takara, 9109) and then incubated with DNase I (Roche, 04716728001) to remove DNA contamination. Total RNA (200 μg) was randomly fragmented into ~200 nucleotides by RNA Fragmentation Buffer (100 mM Tris-HCl, 100 mM ZnCl2 in DEPC-treated water). Protein A magnetic beads (Thermo Fisher, 10002D) were pre-incubated with 5 μg of anti-m6A polyclonal antibody (Synaptic Systems, 202003), and then incubated with fragmented RNA in IP buffer (150 nM NaCl, 10mM Tris-HCl, 0.1% IGEPAL CA-630 in DEPC-treated water). The fragmented RNA binding with Protein A beads-antibody mixture was isolated using RNeasy Mini Kit (QIAGEN, 74106) according to the manufacturer’s instruction.
The library preparation for next-generation sequencing was constructed by TruSeq Stranded mRNA Sample Prep Kit (Illumina, USA) and quantified by BioAnalyzer High Sensitivity DNA chip.

treatment_protocol: HFE145 cells transfected with negative control or 20 nM WTAP siRNAs were infected with or without H. pylori (MOI 1:100) for 24 hours.

value: Normal gastric epithelial cell

tag: sirna

value: siWTAP

value: Co-cultured with H. pylori

tag: antibody


title: p025_promoter_series_Pgk1p_mBC_oBC

description:
Plasmid p025 oBC-mBC subassembly
Escherichia coli (for plasmid amplification)
final_subassembly_oBC_promoters_mBC_triplets_promoter_series.txt

tag: treatment


title: siWTAP.IP

description:
Cells transfected with 20 nM WTAP siRNAs.
m6A-enriched RNA

value: anti-m6A polyclonal antibody


title: MM3_D703_S3


title: p025_promoter_series_UBCp_mBC_oBC


title: liver_WT_d2

city: Braunschweig

first: Andreas

last: Jeron

state: Nierdersachsen

address: Inhoffenstraße 7

department: Viral Immunology

country: Germany

institute: Helmholtz Center for Infection Research

description: Livers from Lm infected wildtype mice on day 2 post infection

hyb_protocol: Hybridization, blocking, staining, washing, was performed acording to GeneChip® Mouse Gene 2.0 ST microarray manufacturer's recommendations

scan_protocol: Scanning was performed acording to GeneChip® Mouse Gene 2.0 ST microarray manufacturer's recommendations

data_processing: MoGene-2_0-st (version 1) data were processed with Transcriptome Analysis Console 4.0 (Thermo Fischer) using RMA algorithm and annotation source file: MoGene-2_0-st-v1.na36.mm10.transcript.csv

label_protocol: Labeling was performed acording to GeneChip® Mouse Gene 2.0 ST microarray manufacturer's recommendations

extract_protocol: RNA from whole liver homogenates was isolated and purified using the RNeasy Mini Kit (Qiagen) and the Rnase-free DNase kit (Qiagen) according to manufacturer recommendations. Per analyzed point in time following Lm infection the RNA concentration of samples from 3-5 WT and IkBNS -/- mice was measured with a photospectrometer (NanoDrop) and equal amounts of RNA from each sample were pooled to obtain a representative sample mix integrating biological variances.

value: Wildtype

tag: infection

value: LM

tag: time

value: 2 Days post LM-infection

value: liver


title: siWTAP.Input

tag: cell line

tag: cell type


title: siWTAP+H. pylori.IP

tag: condition


title: siNC+H. pylori.IP

description:
Cells transfected with negative control were co-cultured with H. pylori (MOI 1:100) for 24 hours.
m6A-enriched RNA

value: siNC


title: BMS2_D705_S5


title: liver_WT_d0

description: Uninfected livers from wildtype mice

source_name: Uninfected livers from wildtype mice

tag: genotype

value: control

value: 0 Days post LM-infection


title: p025_promoter_series_noP_mBC_oBC


title: p025_promoter_series_EEF1A1p_mBC_oBC


title: 210407_HDg1_snRNA-A8

city: Cambridge

first: Myriam

last: Heiman

state: Massachusetts

address: 77 Massachusetts Ave., Bldg 46-4285

department: Brain and Cognitive Sciences

institute: Massachusetts Institute of Technology

description: Human HD grade 1 putamen

library_source: transcriptomic single cell

data_processing:
Basecalls converted to FASTQ with Cellranger v3.1.
FASTQ files were aligned, demultiplexed, and sorted with Cellranger v3.1 using pre-mRNA annotated GRCh38 or mm10 reference.
Genome build: GRCm38/mm10
Supplementary_files_format_and_content: Sparse matrices (MTX) containing unique molecular identifier counts of each gene per cell.
snRNA-seq: Basecalls converted to FASTQ with Cellranger v3.1.
snRNA-seq: FASTQ files were aligned, demultiplexed, and sorted with Cellranger v3.1 using pre-mRNA annotated GRCh38 or mm10 reference.
Genome_build: snRNA-seq: GRCm38/mm10
Supplementary_files_format_and_content: snRNA-seq: Sparse matrices (MTX) containing unique molecular identifier counts of each gene per cell.
TRAP: FASTQs files were aligned to the Mus musculus genome using STAR v2.4.2a. Read counts were generated by STAR using the --quantMode GeneCounts option, along with the default options, except for  --outFilterMismatchNoverLmax, which was set to 0.04 (from 0.3), --outFilterMismatchNmax, which was set at 999 (from 10), --alignIntronMax, which was set to 1000000, and --alignMatesGapMax, which was set to 1000000.
Retro-TRAP: FASTQs files of 50-bp single-end sequencing reads were aligned using STAR 2.4 RNA-Seq aligner. As note, technical replicate data were merged for alignment. The mapped reads were processed with HTSeq to generate gene counts (GENCODE.vM9 version of mouse gene annotation).
Genome_build: TRAP: GRCm38.78, Retro-TRAP: GRCm38.p4
Supplementary_files_format_and_content: TRAP: TSV and TXT files containing raw counts and reads per kilobase mission of each gene per sample.

source_name: Putamen

extract_protocol:
Tissue was homogenized in 700 µL of homogenization buffer (320 mM sucrose, 5 mM CaCl2, 3 mM Mg(CH3COO)2, 10 mM Tris HCl [pH 7.8], 0.1 mM EDTA [pH 8.0], 0.1% NP-40, 1 mM β-mercaptoethanol, and 0.4 U/µL RNase inhibitor) with a 2 mL tissue grinder. Tissue was filtered through a 40 µm cell strainer and mixed with 450 µL solution of 50% OptiPrep density gradient medium, 5 mM CaCl2, 3 mM Mg(CH3COO)2, 10 mM Tris HCl [pH 7.8], 0.1 mM EDTA [pH 8.0], and 1 mM β-mercaptoethanol.  The mixture was then pipetted onto an OptiPrep density gradient containing 750 µL of 30% OptiPrep Solution (134 mM sucrose, 5 mM CaCl2, 3 mM Mg(CH3COO)2, 10 mM Tris HCl [pH 7.8], 0.1 mM EDTA [pH 8.0], 1 mM β-mercaptoethanol, 0.04% NP-40, and 0.17 U/µL RNase Inhibitor) on top of 300 µL of 40% OptiPrep Solution inside a microcentrifuge tube. Nuclei were pelleted at the interface of the OptiPrep density gradient by centrifugation. Nuclei were washed three times with 2% BSA and the nuclear pellet was re-suspended in 100 µL of 2% BSA.
Libraries were constructed according to the 10x Genomics Single Cell Expression v3 User Manual.

value: Putamen

value: HD

tag: tissue id


title: 210407_HDg1_snRNA-A7

description: Human HD grade 1 caudate

source_name: Caudate

value: Caudate


title: MCF-7 H3K4me2 ChIP

description: Chipped

source_name: breast cancer cell line

tag: gender

value: female

value: breast cancer

tag: integrated vector

tag: transfected sirna

value: no

value: 10-8M Estradiol (Sigma)

tag: chip antibody

value: 07-030 (Millipore)


title: BMS3_D706_S6

value: Myeloma Cell Line

value: Malignant plasma cell


title: DNP

city: Zhanjiang

first: Zhencong

last: Li

address: 57 South Renmin Avenue, Xiashan District, Zhanjiang City, Guangdong Province, China

institute: Affiliated Hospital of Guandong Medical University

library_source: genomic single cell

data_processing:
Basecalls performed using CASAVA version 1.8
Umi-tools was utilized to calculate the raw counts and cell whitelist by default parameter
We applied the Seurat [https://satijalab.org/seurat/] package for cell normalization and cell filtering considering the MT percentage, minimum and maximum gene numbers under following criteria:   MT% < 20%;    Cell Gene Number < Median Gene Number * 2;   Cell Gene Number > 200.  PCA and tSNE analysis was used for the single cell to cell relation description. Graphcluster and K-mean was utilized for cell clustering and based on the marker gene achieved some cluster was combined. Wilcox rank sum test was then used for marker gene analysis.
Assembly: GRCh38
Supplementary files format and content: tab-delimited text files include normalized counts for each cell

source_name: Human nucleus pulposus tissue

extract_protocol:
Sample washing with PBS containing 0.04% BSA, the cell pellets were re-suspended in PBS containing 0.04% BSA. Dissociated single cells were then stained with AO/PI for viability assessment using Countstar Fluorescence Cell Analyzer.
Sequencing libraries were conducted using the BD Rhapsody Single-Cell Analysis System (BD Biosciences), following the manufacture's protocol.

value: nucleus pulposus cells

tag: disease state

value: degenerative


title: siNC+H. pylori.Input

value: Normal gastric epithelial cell

value: Co-cultured with H. pylori


title: p025_recloned_complex_mBC_oBC

description:
Plasmid p025 oBC-mBC subassembly
Escherichia coli (for plasmid amplification)
p025_recloned_complex_mBC_oBC_subassembly.txt

value: bacterial cell


title: p025_promoter_series_minP_mBC_oBC


title: liver_KO_d4

description: Livers from Lm infected Nfkbid knockout mice on day 4 post infection

value: Nfkbid knockout

value: LM

value: 4 Days post LM-infection

value: liver


title: liver_KO_d3

description: Livers from Lm infected Nfkbid knockout mice on day 3 post infection

value: 3 Days post LM-infection


title: liver_WT_d3

description: Livers from Lm infected wildtype mice on day 3 post infection

value: Wildtype


title: liver_KO_d2

description: Livers from Lm infected Nfkbid knockout mice on day 2 post infection

value: 2 Days post LM-infection


title: liver_WT_d4

description: Livers from Lm infected wildtype mice on day 4 post infection


title: bulk_MPRA_PolIII_p053_oBC_maps

description:
Bulk MPRA
Escherichia coli (for plasmid amplification)
bulk_MPRA_PolIII_BC_list.txt

data_processing:
Sequencing data was demultiplexed using bcl2fastq. Raw fastq files were processed first by trimming unnecessary cycles from the 3’ end (9 cycles from read 1, 4 cycles from read 2) using seqtk (https://github.com/lh3/seqtk). Forward and reverse barcode reads were joined and error corrected with PEAR (options -v 16 -m 16 -n 16 -t 16). Using custom python and R scripts, successfully assembled barcode reads were combined with UMI reads, barcode/UMI pairs were counted, and the read counts and UMI count per barcode was determined. These barcode count files served as the processed inputs for downstream analysis.
For downstream processing, the identity of barcodes present in each transfected plasmid library (p053 and p054) was first determined by inspecting the distribution of barcode UMI counts from separate libraries directly prepared from the respective plasmids. The count distribution displayed clear bimodal nature, and barcodes in the high count mode (>9 UMIs for p053, >5 UMIs for p054) were retained as valid. Following removal of barcodes present in both libraries (34 out of 193705), we were left with a list of barcodes for expression analysis (59.0k for p053, 134.6k for p054).
For quantifying steady-state expression of linear and circular barcodes, we tallied the UMI counts for all valid barcodes for genomic DNA and RNA derived libraries. DNA UMI counts were well correlated from genomic DNA to plasmid (log-transformed R2 comparison of BC UMI= 0.42, and 0.43 for respectively replicates 1 and 2). Steady-state expression (referred to as “activity”) was defined as the normalized RNA UMI counts over the normalized DNA UMI counts, with normalized UMI counts defined as UMI counts over all UMIs mapping to valid barcodes in the respective libraries. For BC well represented in the library (>50 DNA UMI counts), the activity was >150-fold higher for Tornado barcodes compared to linear barcodes (median activity fold-change 162⨉ in rep1 and 186⨉ in rep2).
Assembly: N/A
Supplementary files format and content: bulk_MPRA_counts_PolIII_oBC_vs_linBC: read and UMI counts for each barcode to the two biological replicates. Column 1: biological replicate id; column 2: Pol III barcode; column 3: barcode type (linear linBC or circular oBC); column 4: UMI counts DNA; column 5: read counts DNA; column 6: UMI counts RNA; column 7: read counts RNA.
Supplementary files format and content: bulk_MPRA_PolIII_BC_list: list of barcodes passing read cutoff thresholds for the mapping of the barcode type (circular barcodes from plasmid p053, linear barcodes from plasmid p054). Column 1: Pol III barcode; column 2: UMI count; column 3: read counts; column 4: corresponding plasmid library; column 6: barcode type.

growth_protocol:
K562 cells were grown in RPMI 1640 medium (ThermoFisher, cat. num. 11875119), supplemented with 10% FBS and 1x Penicillin/streptomycin (ThermoFisher, cat. num. 15140122).
2.75 μL each of plasmids libraries p053 and p054 were mixed to 0.6 μL (0.3 μg) of SBI super piggyBac, and transfected 2.5 M of exponentially growing K562 cells in duplicates using a Nucleofector following manufacturer’s protocol for K562 (kit V4XC-2024, Lonza BioResearch) in duplicates. After two weeks of exponential growth with ⅕ split every two days to allow for dilution of unintegrated plasmids, cells were harvested in exponential phase (<0.75 M/mL), and methanol fixed. Briefly, cells were pelleted at 500 g for 5 min, washed with ice cold PBS to 2 M/mL, pelleted at 500 g for 5 min, resuspended to 15 M/mL, and ice cold methanol was added drop by drop to 80%. Aliquots of 4 M fixed cells were stored at -80C until DNA or RNA extractions.

extract_protocol:
Genomic DNA was extracted from methanol fixed cells using the DNeasy kit (Qiagen), and RNA was extracted from cells using TRIzol LS (Thermo Fisher), following manufacturer’s instructions in both cases.
Amplicon libraries from DNA were generated in two steps of PCR amplification with Kapa HiFi (Roche). For genomic DNA, 500 ng of input was used, and for plasmids (to map barcodes present in both constructs), 3 ng was used. For low-cycle number PCR1, 500 ng of DNA was mixed with 50 μL 2⨉ Kapa HiFi master mix, 5 μL 10 μM o246, 5 μL 10 μM o424, and water to 100 μL. Cycling parameters: 1 min at 95C, and 4 cycles of: 20 s at 98C, 20 s at 60C, 30 s at 72C, followed by 4C hold. Primer o424 contains 10 random Ns to serve as a pseudo-UMI (hereafter referred to as UMIs for brevity) to correct for PCR jackpotting. Reactions were cleaned up with Ampure XP beads (Beckman Coulter) at 1.75⨉, and eluted in 20 μL of 10 mM Tris 8. Illumina adapters and sequencing indices were appended through PCR2, with 4 μL of the eluate from PCR1 taken as input, and 25 μL 2⨉ Kapa HiFi master mix, 0.25 μL 100⨉ SYBr green, 2.5 μL 10 μM o076, 2.5 μL 10 μM indexed primers (DNA rep1: o501, DNA rep2: o502, plasmid p053: o427, plasmid p054: o504), and water to 50 μL. Libraries were amplified with tracking by qPCR with: 1 min at 95C, and cycles up to the qPCR inflection point (typically 15-17 cycles) of: 20 s at 98C, 20 s at 60C, 30 s at 72C. Libraries were then cleaned up with Ampure XP beads at 1.75⨉.
Amplicons libraries for RNA were obtained by first DNase treating the RNA (5 μg RNA, 2 μL TURBO DNase [Thermo Fisher], 2 μL 10⨉ buffer, and waster to 20 μL, incubated at 37C for 30 min, cleaned up with RNA clean & concentrator [Zymo Research], and eluted in 11 Tris 7 10 mM), and taking 1 μg of DNase treated RNA to reverse transcription. Briefly, 2 μL (500 ng/μL) RNA was mixed with 2 μL 1 μM o424, incubated at 65C for 5 min, and placed on ice. 15 μL of reverse transcription master mix was then added (4 μL 5⨉ FS buffer, 1 μL 0.1 M DTT, 1 μL 10 mM dNTP mix, 8 μL water, 1 μL SSIII [Thermo Fisher]), and the reaction incubated at 55C for 60 min, followed by 70C for 15 min. ¼ of the reverse transcription reaction was then directly amplified for PCR1 (37.5 2⨉ Kapa HiFi master mix, 3.75 μL o246, 3.75 μL o076, water to 75 μL), with cycling parameters: 1 min at 95C, and 4 cycles of: 20 s at 98C, 20 s at 60C, 30 s at 72C, followed by 4C hold. Reactions were cleaned up with Ampure XP beads (Beckman Coulter) at 1.75⨉, and eluted in 20 μL of 10 mM Tris 8. PCR2 proceeded as for libraries prepared from plasmids and genomic DNA, with indexing primers o508 and o509 for replicates 1 and 2 respectively, and reactions stopped at inflexion point from qPCR tracking (cycle 7). Libraries were then cleaned up with Ampure XP beads at 1.75⨉.
Read structure: read1, oBC (25 cycles, primer o431); index1, library index (20 cycles, primer o432); read2, oBC reverse (20 cycles, primer o433); index2, UMI (10 cycles, primer o434)
Bulk MPRA


title: liver_KO_d0

description: Uninfected livers from Nfkbid knockout mice

value: control

value: 0 Days post LM-infection


title: siNC.Input

description:
Cells transfected with negative control.
m6A-enriched RNA

value: siNC


title: bulk_MPRA_PolIII_p054_linBC_maps


title: NNP

value: normal


title: soybean_ssDRIP_rep2

status: Public on Dec 10 2022

city: Beijing

first: li

last: kuan

address: zhongguancun

institute: Tsinghua University

data_processing:
library strategy: ssDRIP-seq
Illumina Casava software used for basecalling.
Reads were aligned to the genome of  Glycine max (Zhonghuang13) using bowtie2 version 2.3.3.1.
Duplicated reads were removed by picard version 1.12
Read coverage normalized to 1x sequencing depth (also known as Reads Per Genomic Content (RPGC)) using deeptools version 3.1.2
Assembly: Genome_build: Glycine max (Zhonghuang13)
Supplementary files format and content: bigwig. Each processed bigwig files was used for visualization via IGV.

organism: Glycine max

source_name: leaf

extract_protocol:
Nuclei were isolated from 12-day old soybean leaves using Honda buffer, followed by SDS/proteinase K treatment at 37°C overnight. Genome DNA was extracted by the phenol-chloroform method and precipitated with isopropanol. DNA fragmentation was performed overnight using endonucleases (MboI, MseI, NlaIII, DdeI). DRIP was performed as described in Xu et al., 2017. Briefly, 2 ug of DNA was bound with 30 ug of S9.6 antibody in 1 X binding buffer (10 mM NaPO4 pH 7, 140 mM NaCl, 0.05% Triton X-100) overnight at 4˚C. 50 ul Protein G sepharose beads were added for 4 hours. Bound beads were washed 4 times in binding buffer and elution was performed in elution buffer (50 mM Tris pH 8, 10 mM EDTA, Proteinase K) for 45 min at 55˚C. DNA was purified and sonoicated to 150 bp fragments.
The DNA strands from enriched RNA:DNA hybrids were conducted to single-stranded DNA library construction. The ssDRIP-seq library construction was performed following the instruction manual of VAHTS ssDNA Library Prep Kit for Illumina (Vazyme, ND620).

value: leaf

tag: age

value: 12 days old

tag: infraspecific name

value: Cultivar:Zhonghuang 13

value: wild type


title: soybean_ssDRIP_rep1

tag: tissue


title: ECH2

city: Groningen

first: Anne

middle: de

last: Jong

address: Nijenborgh 7

department: Molecular Genetics

country: Netherlands

institute: university of Groningen

data_processing:
mapping of reads was done using: bowtie2  --local using Escherichia coli K12 as template genome
SAM files were converted to BAM files using SAM tools
BAM files were converted to RPKM values using BED-tools
Count values were normalized using the TMM (Trimmed Median Mean) method in T-REx (doi: 10.1186/s12864-015-1834-4)
Genome_build: Escherichia coli ASM74325v1, NCBI_Assembly:GCA_000743255.1
Genome_build: Staphylococcus aureus ASM75620v1, NCBI_Assembly:GCA_000756205.1
Supplementary_files_format_and_content: Tab delimited text file

source_name: Bacterial

growth_protocol: Cell were grown and treated as described in the attached article

extract_protocol:
RNA was isolated as describe in attached article
Standard library preparation protocol as describe in attached article

tag: strain

tag: sample groupd

value: ECH


title: ECH1


title: EC2

value: EC


title: EC1


title: siNC.IP

value: Normal gastric epithelial cell

tag: sirna

tag: antibody

value: anti-m6A polyclonal antibody


title: TIGIT

city: Seattle

first: Scott

middle: N

last: Furlan

address: 1100 Fairview Ave N

institute: CRD

data_processing:
Data were demultiplexed and counted using Cellranger version 7.00
Resulting fastq files were run using the cellranger multi function with parameters for both gene expression and vdj counting
To achieve similar read depth of gene expression libraries across samples, the cellranger aggregate function was used to downsample reads yielding an expression matrix of gene counts per cell.
Cells meeting the following thresholds were kept for downstream analysis: Unique Molecular Indices per cell > 2500 & < 25000; percent mitochondrial RNA < 15%; cells that exhibited expression of Cd3e/g & Cd8a genes
Assembly: mm10
Supplementary files format and content: Raw counts

source_name: CD8+ T

growth_protocol: Mice on C57BL/6 background (WT or CRBN transgenic) were transplanted with 10 x 10^6 BM with 5 x 10^6  T cells from  genetically matched donors

extract_protocol:
CD8 T cells were FACS sorted from BM (5 mice were pooled per group) at 28 days post-transplant
cDNA was generated using 10X 5' platform per manufacturers instructions

treatment_protocol: Mice were treated with anti-TIGIT (or isotype) from the day of transplant and with lenalidomide (or vehicle) from 14 days post-transplant until harvest

value: BM T cells

tag: treatment

value: TIGIT inhibition + vehicle

tag: timepoint

value: Week 4 post-SCT

value: C57Bl/6 background


title: Len

tag: cell type

value: Isotype control + Lenalidomide


title: Combo

value: TIGIT inhibition + lenalidomide


title: Control

value: Isotype control + vehicle


title: MDA::ER::FOXA1 FOXA2 ChIP

description: Chipped

value: female

tag: cell line

value: breast cancer

value: no

tag: hormone treatment

value: 10-8M Estradiol (Sigma)

value: Antibody Mix:clone 3C10 (Millipore) + ab23630 (Abcam)


title: PIC_14d_049

first: Rob

last: Coppes

address: Ant. Deusinglaan 1

department: Biomedical Sciences of Cells and Systems

institute: University Medical Center Groningen

data_processing:
Base-calling: bcl2fastq software (Illumina Inc.) version 2.20.0.422.
QuantSeq 3′ mRNA-Seq analysis: Bluebee (Lexogen) version 015UG108V0311
Assembly: GRCh38
Supplementary files format and content: Matrix table with raw gene counts.txt for every gene and every sample

molecule: polyA RNA

source_name: PIC-cultured organoid derived from human salivary gland tissue

growth_protocol: SGOs were cultured in Matrigel and PIC for 14 days. RYTN medium consisting of DMEM/F12 [Gibco, 11320-074] containing 1% Pen/Strep antibiotics [Gibco, 15140-122], 1% glutamax [ThermoFisher Scientific, 35050038], 1x N2 [Gibco, 17502-048], 20 ng/ml EGF [Sigma, E9644], 20 ng/ml FGF2 [Peprotech, 100-18B], 10 µg/ml insulin [Sigma, I6634], 1 µM dexamethasone [Sigma, d4902], 10 µM Y-27632 [Abcam, ab120129], 50 ng/ml noggin [Peprotech, 120-10C], 1 µM A8301 [Tocris Bioscience, 2939], and 10% R-spondin1 conditioned medium was added and refreshed every 7 days.

extract_protocol:
RNA was isolated with the RNeasy Mini Kit (Qiagen, 74104) according to manufacturer’s instructions including DNase I treatment
QuantSeq 3’ mRNA-Seq Library Prep Kit (Lexogen)

value: salivary gland organoid

tag: culture condition

value: PIC


title: Matrigel_14d_045

source_name: Matrigel-cultured organoid derived from human salivary gland tissue

value: Matrigel


title: Matrigel_14d_049_2


title: Matrigel_14d_060


title: Matrigel_14d_049


title: PIC_14d_012


title: Matrigel_7d_049


title: PIC_14d_013


title: PIC_14d_045


title: PIC_14d_060


title: Matrigel_7d_060


title: Matrigel_14d_013


title: Matrigel_14d_012


title: Matrigel_7d_045


title: Differentiated 2DF-derived iMACs, cell line K7 (rep 3)

city: Moscow

first: Irina

last: Lyadova

address: 26 Vavilov Street

country: Russia

institute: Koltzov Institute of Developmental Biology of the Russian Academy of Sciences

data_processing:
Trimming: CLC Genomics Workbench 20.0.4 (https://digitalinsights.qiagen.com) was used for read trimming with the following parameters: “quality scores - 0.005; trim ambiguous nucleotides – 2; remove 5’-terminal nucleotides – 1; remove 3’-terminal nucleotides – 1; discard reads below a length of 25”.
Read mapping: The high-quality reads were uniquely mapped using the CLC Genomics Workbench (length fraction = 1 and similarity fraction = 0.95) to the reference genome of H. sapiens.
Readcount obtaining: readcounts were calculated for each gene using CLC Genomics Workbench. Total number of reads mapped on a gene was used as a readcount for the gene.
Assembly: GRCh38
Supplementary files format and content: Raw gene readcounts for each sample.

extract_protocol:
For RNA isolation, the cells were homogenized using syringe and needle or a QIAshredder kit, and RNA was isolated using RNeasy Micro (<5x10^5 cells) or RNAeasy Mini (>5x105 cells) Kits.
RNA concentrations were measured using a Qubit™ RNA HS Assay Kit and RNA integrity number (RIN) was assessed using an RNA 6000 Pico Kit for Bioanalyzer. Only RNA samples with RIN > 8 were used for subsequent library preparation. PolyA RNA was isolated using a NEBNext Poly(A) mRNA Magnetic Isolation Module, containing magnetic Oligo d(T) beads. Transcriptome libraries were constructed using a NEBNext Ultra II RNA Library Prep Kit for Illumina following manufacturer’s instructions. Libraries were prepared with a starting amount of 150 ng and amplified with 15 PCR cycles. RNA was fragmented to the average size of 200nt by incubation at 94 °C for 10 min. Libraries were quantified with a Qubit dsDNA HS Kit and the size distribution was determined with a High Sensitivity DNA Kit for Bioanalyzer.

value: iPSC-derived macrophages

value: 2DF protocol of differentiation, CD14+ iMacs


title: Differentiated 2DF-derived iMACs, cell line iMA (rep 3)

source_name: iMA

value: iMA


title: Differentiated EBS-derived iMACs, cell line K7 (rep 3)

value: EBS protocol of differentiation, CD14+ iMacs


title: Matrigel_7d_012


title: Tissue_025

source_name: fresh human salivary gland tissue

value: salivary gland tissue


title: Tissue_024


title: Tissue_021


title: HS578T_ scRNAseq (GEX+Ab_Capture)

city: Indianapolis

first: Kun

last: Huang

state: IN

address: 410 W 10th street

institute: Indiana University

description: 10x Genomics

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were made using the Cell Ranger software v2.1.1 (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger)
Assembly: hg38
Supplementary files format and content: Tab-separated values files and matrix files

source_name: Breast cancer

extract_protocol:
Freshly collected HS578T cells were flushed with ice-cold HBSSwo (Mg2+/Ca2+ free supplemented with 10mM HEPES, pH 7.2) to remove luminal content. The tissue was inverted and inflated by injection of HBSSwo. Epithelial isolation was performed by incubation of tissue with pre-digestion buffer (HBSSwo, 5 mM EDTA, 5% v/v FCS, 1 mM DTT) in a shaking incubator. The isolated epithelial cells were resuspended in digestion buffer (HBSS with Mg2+/Ca2+ supplemented with 2 mg/ml collagenase type I and 40 U/ml DNase I) and incubated at 37 °C for 30 min. Cells were washed and re-suspended in PBS and stained with a Viability Dye, washed and resuspended in FACS buffer (HBSSwo, 2% v/v FCS, 5mM EDTA). Alive GCs were sorted using a FACS Jazz (Becton Dickinson) according to the presence or absence of mCherry signal using a C57Bl6 mouse as negative control. Goblet cells were washed in HBSS-/- and resuspended in HBSS-/- supplemented with 0.04% w/v BSA.
Library was performed according to the manufacter’s instructions (single cell 3’ v2 protocol, 10x Genomics). Briefly, GCs were resuspended in the master mix and loaded together with partitioning oil and gel beads into the chip to generate the gel bead-in-emulsion (GEM). The poly-A RNA from the cell lysate contained in every single GEM was retrotranscripted to cDNA, which contains an Ilumina R1 primer sequence, Unique Molecular Identifier (UMI) and the 10x Barcode. The pooled barcoded cDNA was then cleaned up with Silane DynaBeads, amplified by PCR and the apropiated sized fragments were selected with SPRIselect reagent for subsequent library construction. During the library construction Ilumina R2 primer sequence, paired-end constructs with P5 and P7 sequences and a sample index were added.

value: Breast cancer

value: Eukaryotic cells

tag: genotype

value: WT & one copy of chrom 17p knockout

value: untreated & Doxorubicin treated


title: Cerebral palsy exercise 1

city: Lincoln

first: Ivan

middle: J

last: Vechetti

state: NE

address: 1700 N. 35th Street

institute: University of Nebraska-Lincoln

data_processing:
Sequencing was performed in the NextSeq 500/550 High Output Kit v2 (51 Cycles using a 75-Cycle Kit) on the Illumina NextSeq 500 platform
Single-end reads for miRNAs were mapped to miRbase version 22 using BowTie2. The alignment was performed by Norgen Biotek using sRNAbench and sRNAtoolbox
Raw counts were acquired with featurecounts and they were used as input for the statistical analysis, using DESeq2
Assembly: Gencode version 22 (hg38)
Supplementary files format and content: tab-delimited text files include raw count values for each Sample

source_name: Extracellular vesicles from plasma

extract_protocol:
Total RNA was isolated from EVs using Qiazol Reagent (Qiagen, Carlsbad, CA, US) and ExoRNAeasy kit (Qiagen) according to the manufacturer’s instructions
Total RNA was sent to Norgen Biotek and picograms of RNA was utilized for library preparation.
library preparation and sequencing was performed by Norgen Biotek using the Small RNA Library Prep Kit (Cat. 63600)

treatment_protocol: Blood was collected before and 30 minutes after a bout of aerobic exercise

value: Extracellular vesicles from plasma

tag: group

value: CP EX

value: aerobic exercise


title: Typically developed exercise 2

value: TD EX


title: Typically developed exercise 1


title: Typically developed exercise 3


title: Cerebral palsy exercise 2


title: Cerebral palsy exercise 3


title: Cerebral palsy control 3

value: CP CON

value: control


title: Matrigel_7d_013

value: salivary gland organoid


title: Typically developed control 3

value: TD CON


title: Cerebral palsy control 1


title: Typically developed control 2


title: Don003_NK_rep1

city: Headington

first: Damien

last: Downes

state: Oxfordshire

address: John Radcliffe Hospital

department: MRC Weatherall Institute of Medicine

country: United Kingdom

institute: The University of Oxford

description: Genome-wide library of transposed fragments

data_processing:
Library Strategy: ATAC-seq
Mapping : bowtie -m 2 --maxIns 350, mm9
For un-mapping reads : trimming adaptors trim_galore -- length 10, combining overlapping R1-R2 reads : flash -m 9 -x 0.125
Combining trimmed+flashed and originally mapped reads, mapping again with  bowtie -m 2 --maxIns 350
Removing read pairs overlapping Blacklisted regions
Removing duplicates : samtools rmdup
Reconstructing sequenced fragments from R1+R2 read pairs ( bedtools bedpe | cut -f 1,2,6) - for visualisation
Pileup of fragments (bedtools genomecov -counts)
Generating bigwig tracks
Genome_build: hg19
Supplementary_files_format_and_content: bigWig : DEEPtools bamCoverage converison of filtered fragments (reconstructed, duplicate-filtered fragments : see above)from bam to bigwig,  (--binSize 10 --normalizeUsingRPKM --minMappingQuality 30)

source_name: NK from Peripheral Blood

extract_protocol:
Fresh blood was sourced as 100 ml whole blood collected from three healthy donors (two males, one female) using EDTA Vacuettes (Becton Dickson). Blood was diluted with PBS and overlaid onto Histopaque-1077 (Sigma) and centrifuged for 30 min at 630 rcf (no brake). Peripheral Blood Mononuclear Cells (PBMCs) were washed in PBS and MACS buffer (PBS, 2 µM EDTA, 0.5% BSA) and sequential stained for selective purification of CD8+, CD14+, CD4+, CD19+, and Natural Killer (NK) cells with Microbead kits (Miltenyi Biotec) following the manufacturer’s instruction for 30 minutes (4 ºC). Stained cell were passed over LS or MS columns before the subsequent staining. For NK cells, non-NK cells were first blocked with a biotin-antibody cocktail before binding to NK microbeads following the manufactures instructions.
Assay for transposition of active chromatin sequencing (ATAC-Seq) was performed as previously published (Buenrostro, 2013) on single cell suspensions. 60000-80000 cells were used per biological replicate. Cells were lysed and nuclei were isolated prior to transposition with Tn5 transposase (Nextera, Illumina) for 30 minutes at 37°C. DNA was purified using a MinElute kit (Qiagen).
Libraries were amplified and barcoded using the NEBNext 2xMastermix (NEB) and the custom primers as published in Buenrostro et al., 2013. ATAC-Seq libraries profiles were visualized using D1000 tape on the Tapestation (Agilent). The libraries were quantified using the universal library quantification kit (KAPA Biosystems).

value: NK, Natural Killer

tag: gender

value: Male


title: Don003_CD8_rep1

source_name: CD8 from Peripheral Blood

value: CD8, Cytotoxic T-cell


title: Don003_CD14_rep2

source_name: CD14 from Peripheral Blood

value: CD14, Monocyte


title: Typically developed control 1


title: Cerebral palsy control 2


title: Don002_CD4_rep2

source_name: CD4 from Peripheral Blood

value: CD4, Helper T-cell


title: Don002_NK_rep2


title: Don002_CD8_rep1


title: Don003_CD8_rep2


title: Don002_CD19_rep2

source_name: CD19 from Peripheral Blood

value: CD19, B-cell


title: Don003_NK_rep2


title: Don002_CD4_rep1


title: Don003_CD4_rep1


title: Don003_CD19_rep1


title: Don002_CD8_rep2


title: Don002_NK_rep1


title: Don003_CD14_rep1


title: Don002_CD19_rep1


title: Don002_CD14_rep1


title: Don002_CD14_rep2


title: Don001_CD8_rep1

value: Female


title: Don001_NK_rep2


title: Don001_CD8_rep2


title: Don001_NK_rep1


title: Don003_CD19_rep2


title: Don001_CD4_rep2


title: Don001_CD4_rep1


title: Don001_CD19_rep1


title: Don001_CD14_rep2


title: Don001_CD19_rep2


title: Don001_CD14_rep1


title: SAH2

organism: Staphylococcus aureus

value: SAH


title: SA1

value: SA


title: SAH1


title: hESC_rep4_B (ATAC-seq)

data_processing:
Mapping : bowtie -m 2 --maxIns 350, mm9
For un-mapping reads : trimming adaptors trim_galore -- length 10, combining overlapping R1-R2 reads : flash -m 9 -x 0.125
Combining trimmed+flashed and originally mapped reads, mapping again with  bowtie -m 2 --maxIns 350
Removing read pairs overlapping Blacklisted regions
Removing duplicates : samtools rmdup
Reconstructing sequenced fragments from R1+R2 read pairs ( bedtools bedpe | cut -f 1,2,6) - for visualisation
Pileup of fragments (bedtools genomecov -counts)
Generating bigwig tracks (ucsctools bedGraphToBigWig)
Genome_build: hg19
Supplementary_files_format_and_content: [bigWig]  un-normalized coverage of filtered fragments (reconstructed, duplicate-filtered fragments : see above),  density of alignments in a moving 300 bp window with an increment of 30  bp

growth_protocol: Human ESC line H1 (H1-hESC; WiCell) was grown on Matrigel (Corning) coated plates in mTeSR1 medium (StemCell technologies). Cells were harvested as a single cell suspension using Accutase (EDM Millipore); ATAC-seq and fixation were carried out in mTeSR1 medium.

extract_protocol:
Assay for transposition of active chromatin sequencing (ATAC-Seq) was performed as previously published (Buenrostro, 2013) on single cell suspensions. 60000-80000 cells were used per biological replicate. Cells were lysed and nuclei were isolated prior to transposition with Tn5 transposase (Nextera, Illumina) for 30 minutes at 37°C. DNA was purified using a MinElute kit (Qiagen).
Libraries were amplified and barcoded using the NEBNext 2xMastermix (NEB) and the custom primers as published in Buenrostro et al., 2013. ATAC-Seq libraries profiles were visualized using D1000 tape on the Tapestation (Agilent). The libraries were quantified using the universal library quantification kit (KAPA Biosystems).

value: Embryonic stem cells


title: hESC_rep1_B (ATAC-seq)


title: hESC_rep2_B (ATAC-seq)


title: hESC_rep4_A (ATAC-seq)


title: SA2


title: hESC_rep2_A (ATAC-seq)


title: hESC_rep3_B (ATAC-seq)


title: hESC_rep1_A (ATAC-seq)


title: mCherry_Kidney_270919

city: London

first: LMS

last: Bioinformatics Core

address: Hammersmith Hospital Campus, Du Cane Road

department: Bioinformatics Core

institute: MRC London Institute of Medical Sciences

data_processing:
The raw 10X single-cell RNA sequencing data were mapped and quantified using the 10X Genomics CellRanger pipeline v3.0.2 against the mouse genome mm10 (mm10-3.0.0 reference provided by 10X Genomics).
Genome_build: mm10
Supplementary_files_format_and_content: CSV file containing normalized counts matrix

source_name: Kidney

growth_protocol: p16-FDR mice were generated containing a mammalian optimised FLP recombinase along with a diphtheria toxin receptor (DTR) that is fused with an mCherry fluorescent reporter.

extract_protocol:
p16-FDR mCherry cells were sorted by FACS prior to being centrifuged and resuspended in complete DMEM to a single cell suspension concentration of 700-1000 cells per microlitre. Cells were partitioned and barcoded using the 10X Genomics Chromium Controller and associated v3 kit components, according to manufacturer’s instructions. The harvested cDNA was assessed and quantified using the Agilent 2100 Bioanalyser and library construction performed using the 10X Single Cell 3' Library Kit v3. The libraries were quality controlled by performing a MiSeq Nano PE100 run followed by deeper sequencing on the HiSeq2500 v4 system.
scRNA-Seq libraries were prepared for sequencing using standard 10X Genomics protocol
10X Single Cell 3' Library Kit v3 (scRNA-Seq)

treatment_protocol: p16-FDR mCherry cells from the kidney were FACS sorted.

value: mCherry+ kidney cells


title: miR-486 2

description: Poly-A enrichment

data_processing:
High-throughput sequencing is transformed to sequenced reads (called Raw Data or Raw Reads) by CASAVA base recognition (Base Calling). Raw data are stored in FASTQ (fq) format files, which contain sequences of reads and corresponding base quality.
Paired-end reads for mRNAs were mapped to mm10 version of the mouse genome using STAR. The alignment was performed by Novogene.
Raw counts were acquired with featurecounts and they were used as input for the statistical analysis, using DESeq2.
Supplementary files format and content: mm10
Supplementary files format and content: Tab-delimited text files include count values for each Sample.

growth_protocol: The skeletal muscle C2C12 (ATCC, CRL-1772) myoblasts were cultured in Dulbecco's Modified Eagles Medium (DMEM) supplemented with 10% Fetal Bovine Serum (FBS), 100 units/ml of Penicillin and 100 µg/ml of Streptomycin until reaching 60-70% confluency.

extract_protocol:
RNA extraction was performed using TRI reagent (Sigma-Aldrich, St. Louis, MO, USA) coupled with further columns-based processing using the Direct-zol Kit (Zymo Research, Irvine, CA, USA).
Total RNA was sent to Novogene for library prep.

treatment_protocol: C2C12 myoblasts were transfected with either a mirVana miR-486 mimic or a scramble miRNA sequence (Life Technologies) used as a negative control. This control sequence does not target any known mature miRNA. Transfection was performed using lipofectamine2000 (Life Technologies), according to the manufacturer's instructions. Shortly, lipofectamine complexes were stabilized in OptiMEM (Gibco) for 5 min and incubated with the miRNA probes, diluted to 25 nM in OptiMEM, for 20 min. Finally, the lipofectamine/probe complexes were added to the cells in a medium without antibiotics. After 6 hours of incubation, fresh media was added, and cells were harvested following 24 hours.

value: Myoblast


title: mCherry_Heart_270919

source_name: Heart

treatment_protocol: p16-FDR mCherry cells collected from the heart were FACS sorted.

value: mCherry+ heart cells


title: Control 3

description: Poly-A enrichment

value: Control scramble miRNA


title: Control 1

description: Poly-A enrichment


title: Control 2

description: Poly-A enrichment


title: miR-486 1

description: Poly-A enrichment


title: hESC_rep3_A (ATAC-seq)


title: mCherry_Lung_2700919

data_processing:
A reference genome was created by adding the sequence of mCherry as another gene to the mouse genome reference mm10 (mm10-3.0.0 reference provided by 10X Genomics).
The 10X CellRanger pipeline v3.0.2 was used with this modified reference genome to align the lung dataset.
Genome_build: mm10
Supplementary_files_format_and_content: CSV file containing normalized counts matrix

source_name: Lung

treatment_protocol: p16-FDR mCherry cells from the lung were FACS sorted.

value: mCherry+ lung cells


title: mCherry_Lung_011019


title: TS543-RNF8_3

city: Singapore

first: Oleg

last: Grinchuk

address: 61 Biopolis Dr, Proteos

country: Singapore

institute: IMCB

data_processing:
RNA-seq data quality was monitored via FASTQC package (https://www.bioinformatics.babraham.ac.uk/projects/fastqc/).
Reads preprocessing was performed by Trimmomatic (version 0.36) with the parameters: LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 MINLEN:50.
Mapping of RNA-seq reads was done using STAR (version 020201, Dobin et al., 2013) with default parameters for RNA-seq data; the human ENSEMBL gene annotation model (GRCh38).
RSEM software (Li and Dewey, 2011)  were used to quantify the gene-level expression.
Genome_build: hg38
Supplementary_files_format_and_content: txt format; TPM values for two experimental conditions with 3 replicates each.

source_name: Cancer stem cells

growth_protocol: The GSCs were cultured in human neural stem cell Maintenance Media (Millipore), 1% Penicillin-Streptomycin (PS) supplemented with EGF and  FGF (20 ng/ml each) to maintain stemness.

extract_protocol:
Three days after treatment, cells were harvested and total RNA were extracted with RNeasy® Micro Kit (Qiagen).
A modified NEBNext RNA Ultra protocol is used to prepare the libraries for mRNA sequencing. The first step is selectively purifying the poly-A containing mRNA molecules using oligo-dT attached magnetic beads. Following purification, the mRNA is fragmented using divalent cations under elevated temperature. Next, the cDNA is synthesized using Reverse transcriptase and random hexamers in a first strand synthesis reaction. Subsequently, the cDNA is converted to double stranded cDNA in a DNA polymerase based reaction. These DNA molecules are then subjected to a series of enzymatic steps for end repair, A-Tailing and ligation with NEB adapters. Finally, these adapter ligated molecules are enriched in a limited cycle PCR with indexed primers and purified using AMPure beads to create final cDNA library for sequencing.

treatment_protocol: TS543 cells were transduced with either  GFP vector or GFP RNF8 overexpression construct.

value: Glioblastoma patient derived cancer stem cell

tag: passage


title: TS543-RNF8_2


title: miR-486 3

description: Poly-A enrichment


title: TS543-RNF8_1


title: TS543-GFP_3

value: GFP


title: TS543-GFP_2


title: mCherry_Heart_011019


title: TS543-GFP_1

treatment_protocol: TS543 cells were transduced with either GFP vector or GFP RNF8 overexpression construct.


title: PB019.GATAGGAG

city: New York

first: Peter

middle: A

last: Sims

state: NY

address: 3960 Broadway, Lasker 203AC

institute: Columbia University

data_processing:
Low-input bulk RNA-seq: Reads were aligned to the mouse reference genome GRCm38 and transcriptome annotation (Gencode vM10) using the STAR aligner with parameters –sjdbOverhang 65 –twopassMode Basic after trimming poly(A)-tails from the 3’-ends. The aligned reads were demultiplexed using the well-identifying barcodes, correcting all single-nucleotide errors. All reads with the same well-identifying barcode, UMI, and gene mapping were collapsed to represent an individual transcript. To correct for sequencing errors in UMIs, we further collapsed UMIs that were within Hamming distance one of another UMI with the same well-identifying barcode and gene. For each 96-well plate, after generating a final list of individual transcripts with unique combinations of well-identifying barcodes, UMIs, and gene mapping, we produced a molecular count matrix for downstream analysis.
Single-nucleus RNA-seq: snRNA-seq data were processed as described previously (Yuan et al., 2018) (code available at: https://github.com/simslab/DropSeqPipeline8). Reads were aligned to the mouse reference genome GRCm38 and transcriptome annotation (Gencode vM10) using the STAR aligner with parameters –sjdbOverhang 65 –twopassMode Basic after trimming poly(A)-tails from the 3’-ends. All reads that uniquely aligned to a given gene body (including introns) were kept for downstream filtering analysis, because a large fraction of nuclear transcripts are unspliced. The aligned reads were demultiplexed using the cell-identifying barcodes, correcting all single-nucleotide errors. All reads with the same cell-identifying barcode, UMI, and gene mapping were collapsed to represent an individual transcript. To correct for sequencing errors in UMIs, we further collapsed UMIs that were within Hamming distance one of another UMI with the same cell-identifying barcode and gene. For each sample, after generating a final list of individual transcripts with unique combinations of cell-identifying barcodes, UMIs, and gene mapping, we produced a molecular count matrix for downstream analysis. We first performed unsupervised clustering on the count matrices using the PhenoGraph (Levine et al., 2015) implementation of Louvain community detection after selection of highly variable genes and construction of a k-nearest neighbors graph as described previously (Levitin et al., 2019). We identified 38 clusters (Supp. Fig. 4a), two of which showed statistically significant co-enrichment of dopamine neuron markers such as Th and Slc6a3 based on the binomial test for expression specificity (Shekhar et al., 2016). We identified six major cell types with sufficient coverage for downstream analysis as shown in the UMAP (Becht et al., 2018) embedding in Figure 3a. The assignment of PhenoGraph clusters into major cell type groupings was based on key marker genes significantly enriched in each cluster, as shown in Supp. Fig. 4b: Oligodendrocytes (Mal, Mag, Mog), Astrocytes (Gja1, Fgfr3, Atp1a2), Microglia (C1qb, Siglech, Csf1r), VGLUT2 neurons (Slc17a6, Cacna2d1, Ntng1), GABA neurons (Slc32a1, Gad1, Gad2), and DA neurons (Th, Slc6a3, En1, Slc10a4). Rare populations with insufficient cell numbers (e.g., mural cells, T cells) or ambiguous neuronal clusters possibly containing multiplets (e.g., Gad1+/Gad2+/Slc17a7+/Th+) were excluded from downstream analysis.
Assembly: GRCm38 / Gencode vM10
Supplementary files format and content: Tab-delimited text files containing count matrix for each cell or sample with sample or well-identifying barcode in the header.

source_name: ventral midbrain

growth_protocol: DATIRES-Cre:Ifngr1fl experimental litters were bred from a cross of DATIRES-Cre/wt;Ifngr1fl/fl x DATwt/wt;Ifngr1fl/fl mice such that Cre+ mice (DATIRES-Cre/wt;Ifngr1fl/fl) could be compared to Cre- littermates with no modification of the Ifngr1 locus (DATwt/wt;Ifngr1fl/fl).

extract_protocol:
Nuclei were prepared from the ventral midbrain of mice at 48 hours after injection of saline or IFN-γ. Mice were sacrificed by cervical dislocation and brains were rapidly extracted and submerged in ice-cold phosphate-buffered saline (PBS). Brains were placed on an ice-cold brain matrix (Zivic Instruments) and separated into 0.5-1.0 mm sections using ice cold razor blades. Ventral midbrain tissue was dissected from slices between approximately -2.5 mm to -3.75 mm AP to Bregma. First, the cortex, hippocampi, and any hypothalamus or white matter ventral to the midbrain were removed. A single horizontal cut was made just dorsal to the rostral linear nucleus and all dorsal tissue was discarded. The remaining tissue containing the SN/VTA was flash frozen on liquid nitrogen and stored at -80°C.  Nuclei preparation and sorting were conducted as described by Krishnaswami et al. (2016) with only minor modifications. Frozen tissue was thawed in nuclear isolation medium 1 (NIM1: 250 mM sucrose, 25 mM KCl, 5 mM MgCl2, 10 mM Tris, pH 8.0) supplemented with 1 mM DTT, 0.2 U/µL SUPERaseIN, 1x EDTA-free Protease Inhibitor Cocktail (Roche), and 0.1% Triton X-100. Tissue was homogenized on ice in a glass-glass dounce homogenizer with 30 gentle strokes of loose and tight clearance pestles. All subsequent purification steps were performed on ice or at 4oC unless otherwise specified. Lysates were spun at 500xg for 5 minutes, resuspended in NIM1 supplemented as above and also with 1% BSA, and passed through a 40µm cell strainer cap. Nuclei were centrifuged again at 500xg for 5 minutes and resuspended in staining buffer (PBS supplemented with 1% BSA, 5 mM MgCl2, 0.2 U/µL SUPERaseIN, 1 µg/mL propidium iodide, and 2 µg/mL mouse anti-NeuN AlexaFluor488). After 30 minutes incubation on ice, nuclei were centrifuged at 500xg for 5 minutes and resuspended in 0.5 mL of FACS buffer (PBS + 0.04% BSA, 0.2 U/µL SUPERaseIN, and 1 µg/mL propidium iodide) for sorting.    Nuclei sorting was conducted on a BD Influx (BD Biosciences) using a 100 μm nozzle at 11.1 psi. Single nuclei were gated first using FSC-H vs. FSC-A and then on propidium iodide fluorescence. NeuN/AlexaFluor488 fluorescence was used to establish NeuN- and NeuN+ gates on the single nuclei population. Nuclei were sorted directly into 96 well plates (for bulk nuclei RNA-seq with 96-well plate, pooled library construction) or into 1.5 mL Eppendorf tubes that had been previously coated with 0.1% BSA in PBS overnight at 37oC (for snRNA-seq).
Low-input bulk RNA-seq: The protocol for plate based, 3’ end unique molecular indicator (UMI)-based RNA sequencing of single cells has been described previously (Snyder et al., 2019) as well as modifications to accommodate ultra-low input samples (Hobson et al., 2022b). See Supplementary File 1 for sequences of all custom primers and oligonucleotides used in this protocol. For neuronal and glial nuclei analysis in Figure 1-2, 100 NeuN- or NeuN+ nuclei were sorted directly into the wells of a 96 well plate containing 3 µL of nuclease-free water containing 1 U/µL SUPERaseIN (sort volume ~3 µL for total volume of ~6 µL). After adding 1.5 µL of 10 µM barcoded RT primer (Integrated DNA Technologies), primer annealing was performed at 72°C for 3 minutes. Reverse transcription was performed by adding 7.5 µL RT mix to each well (2.81 µL of 40% polyethylene glycol 8000, 0.15 µL of 100 mM dNTPs, 3 µL of 5X Maxima H RT Buffer, 0.2 µL of 200 U/µL Maxima H Reverse Transcriptase (ThermoFisher), 0.2 µL of 20 U/µL SUPERaseIN, and 0.15 µL of 100 µM Template Switching Oligo (Integrated DNA Technologies), and 1 µL of nuclease free water). Reverse transcription was performed at 42°C for 90 minutes, followed by 10 cycles of 50°C for 2 minutes, 42°C for 2 minutes, 75°C for 10 minutes, followed by a 4°C hold. Excess primers were removed by adding 2 µL of Exonuclease I mix (1.875U ExoI in water) to each well and incubating at 37°C for 30 minutes, 85°C for 15 minutes, 75°C for 30 seconds, 4°C hold. All wells were pooled into a single 15-ml falcon tubes and cDNA was purified and concentrated using Dynabeads™ MyOne™ Silane beads (ThermoFisher) according to the manufacturer’s instructions. The cDNA was split into duplicate reactions containing 25 µl cDNA, 25 µl 2x HIFI HotStart Ready Mix (Kapa Biosystems), and 0.2 M SMART PCR Primer. PCR was run as follows: 37°C for 30 minutes, 85°C for 15 minutes, 75°C for 30 seconds, 4°C hold. Duplicate reactions were combined and purified using 0.7 volumes AMPure XP beads (Beckman Coulter). The amplified cDNA was visualized on an Agilent 2100 Bioanalyzer and quantified using a Qubit II fluorometer (ThermoFisher). Single-nucleus RNA-seq: NeuN nuclear sorting was conducted as described above. Approximately 70% NeuN+ and 30% NeuN- single nuclei were sorted per sample. Sorted nuclei were placed on ice and immediately submitted to the Columbia Single Cell Analysis Core (Columbia Genome Center) for 10X Genomics Chromium Single Cell 3’ v3 library preparation according to the manufacturer's instructions.
low-input bulk RNA-seq (PLATE-seq) and single-nucleus RNA-seq

treatment_protocol: All surgical procedures were approved by the Institutional Animal Care and Use Committee and the Department of Comparative Medicine at New York State Psychiatric Institute. Mice were anesthetized with 4% isoflurane. Animals were transferred onto a Kopf Stereotaxic apparatus and maintained under isoflurane anesthesia (1–2%). After hair removal and sterilization of the scalp using chlorhexidine and ethanol, a midline incision was made. Bregma and Lambda coordinates were determined, and minor adjustments in head position were made to match the DV coordinates. Saline or IFN-γ was injected at AP –3.2, ML −0.9, and DV −4.4. A small hole was drilled into the skull and 400 nL of 0.9% saline or 0.9% saline + 100 ng/µL recombinant mouse IFN-γ (40 ng IFN-γ total) was injected through a pulled glass pipet using a Nanoject 2000 (Drummond Scientific; 8 pulses of 50 nL). At 5 minutes after injection, the glass pipet was slowly withdrawn over 5 min. After closing the skin with vicryl sutures, mice received 0.5 mL of 0.9% saline i.p. and were allowed to recover for >1 hour before being returned to their home cages.

value: ventral midbrain

value: NeuN+ nuclei

value: wildtype

value: IFNG

tag: time point

value: 72 hrs


title: PB019.CCCATACT


title: PB019.TGGAAAGC


title: BMI Epididymis RNAseq rep5

city: Kunming

first: Jinlong

last: Huo

state: Yunnan

address: Jin-hei

department: Faculty of Animal Science and Technology

institute: Yunnan Agricultural University

data_processing:
PacBio long-reads bam files were processed using SMRT Link v9.0 followed by the IsoSeq v3 pipeline (https://github.com/PacificBiosciences/IsoSeq). IsoSeq long-reads processing steps include circular consensus sequence (CCS) generation using ccs, primer removal and demultiplexing using lima. The above workflow generates full-length non-chimeric (FLNC) reads and non-full-length reads based on the presence of 5′ and 3′ adapters and the presence of poly-A tails.
RNA-seq short reads were trimmed using cutadapt v3.0 by removing adapter sequences and low-quality bases. Trimmed reads were aligned to the Sus scrofa reference genome assembly Sscrofa11.1 using STAR and the featureCounts program from Subread package was used to quantify the RNA-seq datasets.
Genome_build: Sus Scrofa genome (assembly Sscrofa11.1, Breed: Duroc, https://www.ncbi.nlm.nih.gov/assembly/GCF_000003025.6/)
Supplementary_files_format_and_content: PacBio: Full-length non-chimeric (FLNC) reads in fasta format. RNAseq: featureCounts results of gene raw counts.

organism: Sus scrofa

source_name: BMI Epididymis

growth_protocol: BMI boars were raised at Yunnan Agricultural University and were harvested at the age of 12-month old.

extract_protocol:
Total RNA was extracted from five mixed testicles samples and five mixed epididymal samples using TRIzol (Thermo Fisher Scientific, Invitrogen, Catalog #15596018) following the protocol provided by the manufacturer. The RIN (RNA integrity number) value of RNA samples was determined by an Agilent 2100 Bioanalyzer and 1% agarose gel electrophoresis. The quality control step ensures that RIN values are above 7.5 and 28S/18S ratios are greater than or equal to 1.5. The purity and concentration of RNAs were detected with a Nanodrop microspectrophotometer (Thermo Fisher Scientific).
Oligo(dT) magnetic beads were used to enrich RNAs with poly-A tails and a SMARTer cDNA synthesis kit (Takara, Clontech, Catalog #634925) was used to generate full-length cDNA molecules. For Illumina RNAseq, the cDNA fragments were purified with a QiaQuick PCR extraction kit (Qiagen, Venlo, The Netherlands, Catalog # 28106), followed by end repair, base addition, and ligation to sequencing adapters. The ligation products were further size selected by agarose gel electrophoresis and amplified by PCR.

treatment_protocol: No treatment prior to the sample collection.

value: Epididymis

tag: breed

value: Banna mini-pig inbred line (BMI)

tag: age

value: adult


title: PB019.AAGGCTCT

tag: strain

tag: tissue


title: BMI Epididymis RNAseq rep4


title: PB019.TCCGCTTT


title: PB019.ACCCCTAA


title: PB019.GACCAGTT


title: PB019.GTACGACA


title: PB019.GTGTTGGA


title: BMI Epididymis RNAseq rep3


title: BMI Epididymis RNAseq rep2


title: BMI Testis RNAseq rep4

source_name: BMI Testis

value: Testis


title: TSPC-U0126-1

city: Guangzhou

first: Liangliang

last: Xu

state: Yuexiu

address: Guangzhou

institute: Guangzhou University of Chinese Medicine

description:
bioreplicate-1
MRNA190919GZ-UO126-1

data_processing:
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to rn6 whole genome.
Genome_build: Jul.2014 (RGSC 6.0/rn6)
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

source_name: Tendon

extract_protocol:
Total RNAs were harvested using Trizol reagent. The preparation of whole transcriptome libraries and deep sequencing were performed by the Sangon Biotech Corporation (Shanghai, China).
RNA libraries were prepared for sequencing using standdard Illumina protocol.

value: tendon

value: TSPC


title: PB019.GCCTAATG

value: 48 hrs


title: PB019.ACGTGTGA


title: PB019.GGGTATCT


title: TSPC-U0126-2

description:
bioreplicate-2
MRNA190919GZ-UO126-2


title: PB019.AGGTTAGG


title: TSPC-control-2

description:
bioreplicate-2
MRNA190919GZ-DMSO2

value: DMSO


title: BMI Testis RNAseq rep3


title: TSPC-control-1

description:
bioreplicate-1
MRNA190919GZ-DMSO1


title: BMI Testis RNAseq rep2


title: PB019.CTGCCAAT


title: PB019.CTGTGTAC


title: PB019.GAGGACTA


title: BMI Testis RNAseq rep1


title: PB019.CTGAAGCT


title: tubal ectopic pregnancy rep5

city: Shanghai

first: Xiaoya

last: Zhao

state: -- None selected --

address: No. 910, Hengshan Rd

institute: School of Medicine, Shanghai Jiaotong University

data_processing:
Illumina Casava1.8 software used for basecalling.
removing  reads  containing  ploy-N, with  5’ adapter contaminants,  without  3’ adapter or the insert tag, containing ploy A or T or G or C and low quality reads from raw data.
The small RNA tags were mapped to reference sequence by Bowtie-0.12.9
Mapped small RNA tags were used to looking for known miRNA. miRBase20.0 was used as reference, modified software mirdeep2(Friedlander et al., 2011) and srna-tools-cli were used to obtain the potential miRNA and draw the secondary structures.
miRNA expression levels were estimated by TPM (transcript per million) through the following criteria (Zhou et al., 2010):Normalization   formula:   Normalized   expression   =   mapped   readcount/Total reads*1000000
Supplementary files format and content: tab-delimited text files include TPM values for each Sample ...
Supplementary files format and content: Matrix table with raw gene counts for every gene and every sample

source_name: villi

extract_protocol:
placental villi were removed, flash frozen on dry ice, and RNA was harvested using Trizol reagent. NEBNext® Ultra™ RNA Library Prep Kit for Illumina® (#E7530L, NEB, USA) was used with 2 ug of total RNA for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: villi

value: placenta

tag: gestational days


title: BMI Testis RNAseq rep5


title: BMI Epididymis RNAseq rep1