title: Pooled input DNA

city: Brussels

first: Eleonore

last: Maury

address: Avenue Hippocrate, 55

country: Belgium

institute: UCLouvain

description: preadipocytes isolated from omental adipose tissue

data_processing:
Reads were aligned to the human genome (hg38) using Burrows-Wheeler alignment algorithm with default settings. Only reads that passed Illumina’s purity filter, aligned with no more than 2 mismatches, and mapped uniquely to the genome were used in the subsequent analysis. In addition, duplicate reads were removed.
For comparative analysis, standard normalization was achieved by down-sampling the usable number of tags (5 ́-ends of the aligned reads) for each sample in a group (non-obese vs. obese OPAs) to the level of the sample in the group with the fewest usable number of tags. This resulted in an equal number of tags (20,191,577 tags) within each group. The tags were extended in silico using Active Motif software at their 3 ́- ends to a length of 200 bp (average fragment length in the size selected library bp). To identify the density of fragments (extended tags) along the genome, the genome was divided into 32-nt bins and the number of fragments in each bin was determined. This information was stored in bigWig files for display. Peak calling was performed using the MACS 2.1.0 algorithm with a default cutoff of p value 10-7 for narrow peaks and 10-1 for broad peaks. Peak filtering was performed by removing false ChIP-Seq peaks as defined within the ENCODE blacklist. This process allowed to determine the significant enrichments in the ChIP/IP data file when compared to the Input data file (~background). Then, known motifs were identified with the findMotifsGenome program of the HOMER package using default parameters and input sequences comprising +/- 100 bp from the center of the top 1000 peaks. Identification of peaks with statistical enrichment over input across conditions was performed using DESeq2 analysis69. The threshold for the number of tags that determines a valid peak was selected for a FDR-adjusted p value of <0.1. This analysis enabled to detect a differential BMAL1 binding genome-wide between non-obese and obese OPAs.
Genome_build: Hg38
Supplementary_files_format_and_content: BigWig files have been generated by Active Motif’s Analysis Program. Supplementary files format and content: bigwig files contain tracks for visualization on the UCSC genome browser

source_name: pooled input (chromatin from 6 patients above)

growth_protocol: OPAs were obtained and cultured as previously described (Maury et al., 2007). Briefly, adipose tissue was finely minced in Krebs buffer, pH 7.4, containing 2% BSA [Krebs-albumin buffer (KRAB)]. The blood vessels were carefully dissected under binocular microscope and discarded. Minced tissue was incubated for ~30 min in a shaking water bath at 37 C in KRAB with collagenase (type 1, Thermo Fisher Scientific, # 17100-017) and filtered through a nylon mesh (200  m). Digestion was stopped by adding DMEM medium (Thermo Fisher Scientific, # 31885023) supplemented with 10% FBS (Thermo Fisher Scientific, # 10500064). The pelleted cells were separated from the adipocyte layer by centrifugation, washed in DMEM with 10% FBS, and this step was repeated three times. After treatment with an erythrocyte lysis buffer, the cells were further processed through a 70-  m cell strainer, washed once in DMEM, and cultured as described (Maury et al, 2007). At ~90% confluency, the cells were trypsinized, aliquoted and the seed working stocks were cryopreserved for subsequent cultures followed by the ChIP experiments.

extract_protocol:
Lysates were clarified from sonicated nuclei.  30 μg of sheared chromatin (300-500 bp) was used per ChIP reaction. Protein-DNA complexes were incubated with 8 μl antibody against BMAL1 (Abcam, #Ab3350).
Illumina sequencing libraries were prepared from ChIP and pooled input chromatin DNAs. Sequencing was generated using 75-bp single-end reads on an Illumina NextSeq 500 instrument to a depth of > 30 million reads.

treatment_protocol: We defined the OPAs as cells positive for Platelet-Derived Growth Factor Receptor alpha (PDGFRa+ or Cluster of Differentiation 140a+, CD140a+) and negative for markers of leucocytes or endothelial cells (CD45 or -CD31-) and determined the expression of CD9 by the following approach. Approximately 97.5 %- 98 % of the cells were PDGFRa+ as determined by flow cytometry analysis using Alexa Fluor 647 mouse anti-human CD140a (BD Biosciences, #562798). Detection of varying amount of CD9 expression in PDGFRa+ OPAs was performed by simultaneous staining with Alexa Fluor 647 mouse anti-human CD140a and CD9-PE antibodies (BD Biosciences, #555372), using a FACScalibur cell analyzer, CellQuest and FlowJo Software (BD Biosciences). No macrophages or endothelial cells, which have a morphologic appearance distinct from OPAs, were seen in cultures, an observation that was further validated by the absence of CD31+ and CD45+ cells by flow cytometry using mouse anti-human CD45-FITC (Immunotools, #21810453X2) and mouse anti-human CD31-FITC antibodies (Miltenyi Biotec, #130-117-539) separately in PDGFRa+ cells. Additionally, while detected in undigested omental adipose tissue, CD31 and CD45 gene expression was undetectable in paired OPAs (i.e, from same patient), confirming the negligible amount of endothelial cells and leucocytes.

value: adipose precursor cells, omental preadipocytes (OPAs)

tag: passages

tag: patient diagnosis

value: pooled sample with DNA from both obese and non-obese patients


title: H3K9me3 in vehicle-treated control rat rep3

status: Public on Jan 29 2021

type: SRA

city: Fujusawa

first: Satoru

last: Matsuda

state: Kanagawa

address: 26-1, Muraoka-Higashi 2-Chome

country: Japan

institute: Takeda Pharmaceutical Company Limited

description:
30_Control-3_H3K9me3_rn-dm_i85
HistonePath_rn.txt.gz

library_source: genomic

data_processing:
Sequences: Standard Illumina software base-calling and quality-control filtering was applied
Sequences (75 nt reads, single end) were aligned to the mouse genome (mm10) or rat genome (rn5) using the BWA algorithm (v0.7.12, default parameters).
Aligns were extended in silico at their 3’-ends to a length of 200 bp, which is the average genomic fragment length in the size-selected library, and assigned to 32-nt bins along the genome. The resulting histograms (genomic “signal maps”) were stored in bigWIG files.
Peak locations were determined using the MACS algorithm (v2.1.0) with a cutoff of pvalue = 1E-7.
Signal maps and peak locations were used as input data to Active Motifs proprietary analysis program, which creates excel tables containing detailed information on sample comparison, peak metrics, peak locations and gene annotations
Genome_build: rn5 or mm10
Supplementary_files_format_and_content: Peak metrics, peak locations and gene annotations in text format

molecule: genomic DNA

organism: Rattus norvegicus

source_name: Brain cortex

extract_protocol:
Brain cortex tissues were removed, and flash frozen on dry ice. Tissues were fixed with 1% formaldehyde for 15 min and quenched with 0.125 M glycine.  Lysates were sonicated with a microtip sonicator to shear the DNA. Genomic DNA (Input) was prepared by treating aliquots of chromatin with RNase, proteinase K and heat for de-crosslinking, followed by ethanol precipitation. For each ChIP reaction, 30 ug of chromatin was precleared with protein A agarose beads (Invitrogen). ChIP reaction was set up using precleared chromatin and antibody and incubated overnight at 4 C. Protein A agarose beads were added and incubation at 4 C was continued for another 3 hr. Immune complexes were washed, eluted from the beads with SDS buffer, and subjected to RNase and proteinase K treatment. Crosslinks were reversed by incubation overnight at 65 C, and ChIP DNA was purified by phenol-chloroform extraction and ethanol precipitation.
Illumina sequencing libraries were prepared from the ChIP and Input DNAs by the standard consecutive enzymatic steps of end-polishing, dA-addition, and adaptor ligation. After a final PCR amplification step, the resulting DNA libraries were quantified and sequenced on NextSeq 500.

tag: strain

value: Sprague-Dawley

tag: disease state

value: healthy

tag: treatment

value: vehicle

tag: chip antibody


title: Non-Obese3

organism: Homo sapiens

source_name: preadipocytes from non-obese patient 3

tag: cell type

value: non-obese


title: H3K9me3 in vehicle-treated control rat rep2

description:
29_Control-2_H3K9me3_rn-dm_i84
HistonePath_rn.txt.gz


title: H3K9Ac in TAK-418-treated VPA rat rep3

description:
27_Model-CpdH-3_H3K9Ac_rn-dm_i85
HistonePath_rn.txt.gz

value: prenatal VPA injection


title: Mouse 2 CC

city: London

first: Sanjay

middle: Padmakar

last: Khadayate

address: Du Cane Road

country: United Kingdom

institute: MRC London Institute of Medical Sciences

description:
Brooks_InVivo_RAW.csv
M2_2

library_source: transcriptomic

data_processing:
Raw reads were aligned and feature counts extracted using the STAR aligner.
reference genome build:hg38
Supplementary_files_format_and_content: csv files containing unfiltered raw counts drom the STAR aligner

molecule: polyA RNA

source_name: Invasive cells from Corpus callosum of mouse 2

extract_protocol:
RNA was isolated using the RNAeasy Plus Micro Kit (Qiagen)
For RNA-seq of FACS-sorted tumour cells, dsDNA libraries were prepared according to the Smart-seq2 protocol from FAC-sorted GFP+ cells (RIN >8) (Picelli et al., 2014). Next-generation sequencing libraries were prepared from 1ng dsDNA using the Nextera XT DNA library preparation kit (Illumina) and indexed using the Nextera XT index Kit (illumina).

value: Patient-derived Glioma stem cell line (G144)

tag: host strain

value: CD-1 Nude (mouse)

tag: cell line

value: Patient derived line G144 expressing luciferase-GFP

tag: patient derived xenograft

value: 6-8 week old female CD-1 nude mice underwent stereotactic implantation of 1x105 GFP-luciferase expressing GSCs (anteroposterior 0, mediolateral -2.5, dorsoventral -3.5). Tumour growth was monitored using an IVIS Spectrum in vivo imaging system (Perkin Elmer). 10 minutes following i.p. D-luciferin (120 mg/kg, Intrace medical) bioluminescent images were acquired under isoflurane anaesthesia. Tumour size was quantified by calculating total flux (photons/s/cm2) using Living Image software (Xenogen, Caliper Life Sciences). Animals were sacrificed when they showed signs of distress or weight loss.

tag: facs sorting of tumour cells

value: Brains were harvested and tumour regions corresponding to tumour bulk, invaded striatum and corpus callosum were micro-dissected under fluorescent guidance in ice cold HEPES buffered HBSS. Micro-dissected regions were dissociated by incubating with papain (20 units/ml) DNase (0.005%) for 30min at 37°C (Worthington). Samples were triturated in HEPES buffered EBSS to fully dissociate the tissue and centrifuged (3 min, 300xg). Digestion was terminated by trituration in EBSS ovomucoid inhibitor (10mg/ml), albumin (10mg/ml) and DNase (0.005%). Following centrifugation (3 min, 300xg), cells were resuspended in 400 μL FACs sorting buffer containing 2.5% RNAsin and 1/10000 DAPI.

tag: culture conditions


title: Non-Obese2

source_name: preadipocytes from non-obese patient 2


title: H3K9Ac in TAK-418-treated VPA rat rep2

description:
26_Model-CpdH-2_H3K9Ac_rn-dm_i84
HistonePath_rn.txt.gz


title: Mouse 2 Bulk

description:
Brooks_InVivo_RAW.csv
M2_1

source_name: Tumour bulk cells from Mouse 2


title: H3K9Ac in TAK-418-treated VPA rat rep1

description:
25_Model-CpdH-1_H3K9Ac_rn-dm_i82
HistonePath_rn.txt.gz


title: H3K9Ac in vehicle-treated VPA rat rep3

description:
24_Model-Veh-3_H3K9Ac_rn-dm_i81
HistonePath_rn.txt.gz


title: Mouse 1 Bulk

description:
Brooks_InVivo_RAW.csv
M1_1

source_name: Tumour bulk cells from Mouse 1


title: Non-Obese1

source_name: preadipocytes from non-obese patient 1


title: ObeseClassIII3

source_name: preadipocytes from patient 3 (obese Class III)

value: obese Class III


title: H3K9Ac in vehicle-treated VPA rat rep2

description:
23_Model-Veh-2_H3K9Ac_rn-dm_i80
HistonePath_rn.txt.gz


title: H3K9me3 in TAK-418-treated VPA rat rep2

description:
35_Model-CpdH-2_H3K9me3_rn-dm_i90
HistonePath_rn.txt.gz


title: H3K9Ac in vehicle-treated control rat rep3

description:
21_Control-3_H3K9Ac_rn-dm_i77
HistonePath_rn.txt.gz


title: ObeseClassIII2

source_name: preadipocytes from patient 2 (obese Class III)


title: H3K9Ac in vehicle-treated VPA rat rep1

description:
22_Model-Veh-1_H3K9Ac_rn-dm_i78
HistonePath_rn.txt.gz


title: H3K9Ac in vehicle-treated control rat rep2

description:
20_Control-2_H3K9Ac_rn-dm_i76
HistonePath_rn.txt.gz


title: A375_Ago2 RIP-Seq 2

city: St. Petersburg

first: Bongyong

last: Lee

state: Florida

address: 501 6th Ave S

country: USA

institute: The Johns Hopkins University

data_processing:
For Ago2 RIP-seq, Reads were mapped to hg19 via Bowtie2
The peakes were called by MACS2/Homer using all the Bowtie alignments
MACS2 results were reported
For RNA-seq, Illumina Casava1.7 software used for basecalling.
Sequenced reads were mapped using tophat2 v2.1.1, with --b2-sensitive --no-novel-juncs options, mapped reads were filtered by quality >10.
Reads Per Kilobase of transcript per Megabase of library size (RPKM) were calculated in Partek GenomicSuite 6.0.
human-only:  filtered by mm10, then mapped to hg19
mouse-only:  filtered by hg19, then mapped to mm10
Genome_build: hg19; mm10
Supplementary_files_format_and_content: tab-delimited text files include reads and RPKM values for each Sample

molecule: total RNA

growth_protocol: A375 (ATCC CRL-1619), A375/211 (MIR211 overexpressing cells, A375/211/PDK4 (MIR211 and PDK4 overexpressing cells) were grown in Complete Tu Medium containing a 4:1 mixture of MCDB-153 medium with 1.5 g/L sodium bicarbonate and Leibovitz's L-15 medium with two mM L-glutamine, 2% FBS, and 1.68 mM CaCl2 (42). The cells were grown in a humidified incubator at 37°C, 5% CO2.

extract_protocol:
Xenografts were pulverized and RNAs were extracted. For Ago2 RIP-seq, RNAs were extracted with Trizol from immunocomplex on beads.
Ago2 RIP-seq RNAs were subjected to double strand cDNA synthesis using an Ovation RNA-Seq System V2 (NuGen Technologies, Inc., San Carlos, CA). High-throughput sequencing libraries were constructed using the TruSeq ChIP Sample preparation kit (Illumina, San Diego, CA) and sequenced on a HiSeq 2500 (Illumina) with 50 bp read lengths.
RNA-Seq libraries were prepared using Truseq Stranded Total RNA Library preparation kit (Illumina, San Diego, CA) with 1 ug of total RNA.

treatment_protocol: 5 million cells were injected to SCID mice and xenografts were excised after 14 days.

tag: cell line background

value: melanoma cell line

tag: genotype/variation

value: overexpressing MIR211


title: A375_Ago2 RIP-Seq 1


title: ObeseClassIII1

source_name: preadipocytes from patient 1 (obese Class III)


title: H3K9Ac in vehicle-treated control rat rep1

description:
19_Control-1_H3K9Ac_rn-dm_i75
HistonePath_rn.txt.gz


title: H3K4me3 in TAK-418-treated VPA rat rep3

description:
18_Model-CpdH-3_H3K4me3_rn-dm_i74
HistonePath_rn.txt.gz


title: H3K9me3 in vehicle-treated control rat rep1

description:
28_Control-1_H3K9me3_rn-dm_i82
HistonePath_rn.txt.gz


title: H3K4me3 in TAK-418-treated VPA rat rep1

description:
16_Model-CpdH-1_H3K4me3_rn-dm_i72
HistonePath_rn.txt.gz


title: Mouse 1 CC

description:
Brooks_InVivo_RAW.csv
M1_2

source_name: Invasive cells from Corpus callosum of mouse 1


title: HIF1A-/- HCT116, hypoxia, replicate 2

city: Aurora

first: Matthew

middle: D

last: Galbraith

state: CO

department: Department of Pharmacology; Linda Crnic Institute for Down Syndrome

institute: University of Colorado Anschutz Medical Campus

description:
nascent RNA (PRO-seq) from HCT116 cells
HIF1Ako_HYPOX-DMSO.merged.pos-strand.norm1e6.frag-given.tbp-all.bedGraph.gz
HIF1Ako_HYPOX-DMSO.merged.neg-strand.norm1e6.frag-given.tbp-all.bedGraph.gz
PROseq_Hypoxia90_TSS_counts.txt
PROseq_Hypoxia90_body_counts.txt

data_processing:
Signal processing, base-calling, and quality scoring was carried out automatically by the NextSeq 500 RTA software (v2.4.11 Illumina).
Demultiplexing and conversion to FASTQ format was carried out using bcl2fastq (v2.20.0.422, Illumina).
FASTQC (v0.11.5) was used to assess sequence data quality.
fastq files from 2 separate lanes were merged for each sample to obtain sufficient read depth.
Trimming and filtering of low-quality bases/reads were performed using BBDUK from BBTools (v37.99) and FASTQ-MCF from EAUtils (v1.05).
FastQ Screen (v0.9.1) was used to check for common sequencing contaminants.
Alignment to the Human reference genome (GRCh37/hg19) was carried out using Hisat2 (v2.1.0) in unpaired, no-spliced-alignment mode with a GRCh37/hg19 index.
Alignments were sorted and filtered for mapping quality (MAPQ>10) using Samtools (v1.5).
Gene-level count data for transcription start site (TSS, -50 to +500) and gene body (+501 to end) regions were obtained using featureCounts from the Subread package (v1.6.2) with custom annotation files for single unique TSS and gene body regions per gene.
Genomic coverage tracks for positive and negative DNA strands were generated as bedGraph files using HOMER from merged replicate bam files created by first downsampling to match the lower total mapped filter alignments to ensure equal contributions from each replicate.
Genome_build: GRCh37/hg19
Supplementary_files_format_and_content: Tab-delimited text files containing per gene sense-strand tag counts for unique TSS or gene body regions for each sample; bedGraph files with genomic coverage tracks.

source_name: HIF1A knockout HCT116 cells

growth_protocol: HCT116 cells were cultivated in McCoy's 5A medium supplemented with 10% fetal bovine serum and 1% antibiotic-antimycotic. Cells were plated the day before treatment and maintained in a humidified atmosphere with 5% CO2 at 37°C.

extract_protocol:
Nuclear run-on was performed as described previously in Steinparzer et al. Mol.Cell 2019 (DOI: 10.1016/j.molcel.2019.07.034 PMID: 31495563) and based on Mahat et al. Nat Protoc. 2016 (DOI: 10.1038/nprot.2016.086 PMID: 27442863)
PRO-seq library construction was performed as described previously in Steinparzer et al. Mol.Cell 2019 (DOI: 10.1016/j.molcel.2019.07.034 PMID: 31495563) and based on Mahat et al. Nat Protoc. 2016 (DOI: 10.1038/nprot.2016.086 PMID: 27442863)
PRO-seq

treatment_protocol: Cells were maintained in normoxia (as above) or exposed to hypoxic conditions by incubation in a humidified atmosphere containing 1% O2, 5% CO2, and 94% N2 for 90min ± addition of DMSO (vehicle) or 10µM 3MB-PP1.

value: HIF1A-/- HCT116

value: colorectal carcinoma

value: Hypoxia (1% O2 90min) + DMSO

tag: library type

value: strand-specific


title: A375/211_Ago2 RIP-Seq 2

source_name: A375 cells


title: A375/211_Ago2 RIP-Seq 1


title: wild-type HCT116, hypoxia, replicate 1

description:
nascent RNA (PRO-seq) from HCT116 cells
WT_HYPOX-DMSO.merged.pos-strand.norm1e6.frag-given.tbp-all.bedGraph.gz
WT_HYPOX-DMSO.merged.neg-strand.norm1e6.frag-given.tbp-all.bedGraph.gz
PROseq_Hypoxia90_TSS_counts.txt
PROseq_Hypoxia90_body_counts.txt

source_name: wild-type HCT116 cells

value: Wild-type HCT116


title: H3K4me3 in vehicle-treated control rat rep3

description:
12_Control-3_H3K4me3_rn-dm_i68
HistonePath_rn.txt.gz

value: Sprague-Dawley

value: healthy


title: A375/211_Ago2 RIP-Seq input


title: H3K4me3 in vehicle-treated VPA rat rep2

description:
14_Model-Veh-2_H3K4me3_rn-dm_i70
HistonePath_rn.txt.gz

value: vehicle


title: H3K4me3 in vehicle-treated VPA rat rep1

description:
13_Model-Veh-1_H3K4me3_rn-dm_i69
HistonePath_rn.txt.gz

value: prenatal VPA injection


title: HIF1A-/- HCT116, normoxia, replicate 1

description:
nascent RNA (PRO-seq) from HCT116 cells
HIF1Ako_DMSO.merged.pos-strand.norm1e6.frag-given.tbp-all.bedGraph.gz
HIF1Ako_DMSO.merged.neg-strand.norm1e6.frag-given.tbp-all.bedGraph.gz
PROseq_Hypoxia90_TSS_counts.txt
PROseq_Hypoxia90_body_counts.txt

value: Normoxia + DMSO


title: HIF1A-/- HCT116, normoxia, replicate 2


title: H3K4me3 in vehicle-treated control rat rep1

description:
10_Control-1_H3K4me3_rn-dm_i95
HistonePath_rn.txt.gz


title: Fetal gonads, sample 1

city: Davis

first: Pablo

middle: Juan

last: Ross

state: California

address: One Shields Ave

department: Animal Science

institute: University of California, Davis

data_processing:
Demultiplexing of raw reads and mapping to the bovine reference genome (ARS-UCD1.2.96) was performed using the Cell Ranger software (v3.0.2). Reads were filtered and counted through the Cell Ranger Count pipeline.
The R package Seurat (v3.1.0.9003) was used for sample integration, quality control, and secondary analyses. Integration and normalization of the two replicates was performed by identifying “anchors” across datasets. Dimensionality reduction was performed by uniform manifold approximation and projection (UMAP) and differential gene expression was performed using non-parametric Wilcoxon rank sum test, which is part of the standard Seurat pipeline.
Genome_build: ARS-UCD1.2
Supplementary_files_format_and_content: HDF5 compressed sparse column format matrix of read counts containing information such as metadata, features, cell-barcodes and UMI.

organism: Bos taurus

source_name: Fetal gonads

extract_protocol:
Slow-frozen gonads were thawed at 37°C in a water bath,  rinsed in ice-cold PBS and minced into ~0.1 mm pieces. Minced tissue was then incubated in Collagenase Type IV (800U/mL) (LS004186, Worthington) supplemented with DNase I (6U/mL) (04536282001, Roche Applied Science) at 37°C in a prewarmed orbital shaker for 10 min at 150 rpm. After dissociation, the cell suspension was filtered and dead cells were removed by magnetic bead sorting using a Dead Cell Removal Kit (13009010, Miltenyi) following the manufacturer’s protocol. Viable cells resuspended in 0.4% BSA were submitted on ice to the UC Davis DNA Technologies & Expression Analysis Core for library preparation and sequencing. Cells were captured using a 10x Chromium controller targeting 10,000 cells per sample.
Libraries were prepared using the Chromium Single Cell 3’ v2 chemistry for sample 1 and v3 chemistry for sample 2 according to the manufacturer’s protocol.

tag: fetus crown-rump length

value: 4 cm

tag: tissue

value: Fetal gonads

tag: gonads size

value: 3 x 1 mm

tag: developmental stage

value: ~day 50 of development


title: Fetal gonads, sample 2


title: H3K4me3 in vehicle-treated control rat rep2

description:
11_Control-2_H3K4me3_rn-dm_i96
HistonePath_rn.txt.gz

tag: strain


title: H3K4me3 in TAK-418-treated VPA rat rep2

description:
17_Model-CpdH-2_H3K4me3_rn-dm_i73
HistonePath_rn.txt.gz

tag: disease state


title: A375/211/PDK4 xenografts_RNA-Seq

description:
A375-211-PDK4-Xeno_S9_L001
processed data file: A375_A375-211_A375-211-PDK4_xeno_*.txt

source_name: A375/211/PDK4 xenografts

value: overexpressing MIR212 and PDK4

value: SCID

tag: sample type

value: xenografts


title: C1 iPSC, 8w, rep2

city: Berlin

first: Petar

last: Glazar

address: Robert Rössle Str. 10 (H. 87)

department: Systems Biology of Gene Regulatory Elements

country: Germany

institute: Max Delbrück Center for Molecular Medicine

data_processing:
Illumina base calling, demultiplexing and FASTQ conversion with bcl2fastq v2.20.0
alignment to GRCh38 genome using STAR v2.6.0c
reads counted on GENCODE 27 gene models with htseq-count 0.9.1
Genome_build: GRCh38
Supplementary_files_format_and_content: htseq-count output, two-column text file with Ensembl gene identifier and read count columns

source_name: dopaminergic neuronal culture

extract_protocol:
RNeasy Mini Kit (Qiagen)
TruSeq RNA Library Prep Kit

tag: status

value: control

tag: timepoint

value: 8 weeks


title: C1 iPSC, 8w, rep3


title: A375_Ago2 RIP-Seq input

tag: cell type

value: melanoma cell line

value: overexpressing MIR211


title: wild-type HCT116, normoxia, replicate 2

description:
nascent RNA (PRO-seq) from HCT116 cells
WT_DMSO.merged.pos-strand.norm1e6.frag-given.tbp-all.bedGraph.gz
WT_DMSO.merged.neg-strand.norm1e6.frag-given.tbp-all.bedGraph.gz
PROseq_Hypoxia90_TSS_counts.txt
PROseq_Hypoxia90_body_counts.txt

tag: cell line

value: Wild-type HCT116

value: colorectal carcinoma

tag: treatment

value: strand-specific


title: A375/211 xenografts_RNA-Seq

description:
211-Tumor-RNA_S25_L008
processed data file: A375_A375-211_A375-211-PDK4_xeno_*.txt

source_name: A375/211 xenografts

tag: host strain


title: C1 iPSC, 8w, rep1


title: C1 iPSC, 4w, rep3

value: 4 weeks


title: wild-type HCT116, hypoxia, replicate 2

value: Hypoxia (1% O2 90min) + DMSO


title: H3K27me3 in TAK-418-treated VPA rat rep2

description:
08_Model-CpdH-2_H3K27me3_rn-dm_i82
HistonePath_rn.txt.gz

source_name: Brain cortex

value: Sprague-Dawley

value: prenatal VPA injection

tag: chip antibody


title: H3K4me3 in vehicle-treated VPA rat rep3

description:
15_Model-Veh-3_H3K4me3_rn-dm_i71
HistonePath_rn.txt.gz

value: vehicle


title: HIF1A-/- HCT116, hypoxia, replicate 1

value: HIF1A-/- HCT116


title: wild-type HCT116, normoxia, replicate 1

value: Normoxia + DMSO


title: NR012: Nonresponder_012

type: RNA

city: Durham

first: Nicholas

last: DeVito

state: NC

address: 2301 Erwin Road

institute: Duke University

description:
Sample acquisition: Archival melanoma specimens of patients treated with anti-PD-1 (either pembrolizumab or nivolumab) monotherapy were obtained via an ongoing clinical trial (NCT02694965). After verification by a board-certified pathologist, microdissection was performed if deemed necessary.
de-identified patient samples were labeled according to response and time of sample acquisition

hyb_protocol: Agilent 2100 Bioanalyzer (Agilent) and NanoDrop (ThermoFisher) were used to determine the purity and concentration of the RNA. A DV300 Agilent Bioanalyzer smear analysis was used to determine the percentage of fragments greater than 300 nucleotides to determine the starting amount of RNA.   Gene expression codesets (designed and produced by NanoString Technologies, Seattle, WA), hybridization buffer and total RNA were hybridized in a thermocycler for 16 hours at 67 °C prior to being processed in the nCounter Max Prep Station following the NanoString manual, MAN-10056-02 for gene expression assays.  Data collection using the nCounter Digital Analyzer was performed by following the NanoString manual, MAN-C0035-07 using the maximum field of view setting.

data_processing: Samples were analyzed on a NanoString nCounter Max system using the nCounter Vantage 3D Human Wnt Pathways Panel (N2_WNT_Pth_v1.0), which is comprised of Wnt ligands, receptors, regulators, gene targets, and other pathway components with the addition of a custom gene panel composed of cytolytic T cell markers and IDO1.  Analysis was performed using the nSolver software.

source_name: Melanoma

extract_protocol: Two-to-three 10 mM FFPE scrolls were collected from FFPE sectioning of biopsies taken prior to treatment, and RNA extraction was performed with using RSC FFPE RNA extraction kit (Promega) according to manufacturer`s instruction.

tag: phenotype

value: Nonresponder

tag: age

tag: disease site

value: Parotid gland metastasis

tag: gender

value: M

value: Melanoma


title: S2_corr organoids, 90 days, rep2

source_name: iPSC-derived cerebral organoids

extract_protocol:
TRIzol
TruSeq Stranded Total LT Sample Prep Kit (Illumina)

value: corrected

value: 90 days


title: C1 iPSC, 4w, rep2

value: control


title: H3K27me3 in TAK-418-treated VPA rat rep1

description:
07_Model-CpdH-1_H3K27me3_rn-dm_i81
HistonePath_rn.txt.gz


title: C1 iPSC, 4w, rep1


title: A375 xenografts_RNA-Seq

description:
Control-Tumor-RNA_S24_L008
processed data file: A375_A375-211_A375-211-PDK4_xeno_*.txt

source_name: A375 xenografts

tag: cell line background

value: melanoma cell line

value: SCID

value: xenografts


title: H3K27me3 in vehicle-treated VPA rat rep3

description:
06_Model-Veh-3_H3K27me3_rn-dm_i80
HistonePath_rn.txt.gz


title: S2_corr organoids, 90 days, rep1


title: H3K27me3 in TAK-418-treated VPA rat rep3

description:
09_Model-CpdH-3_H3K27me3_rn-dm_i84
HistonePath_rn.txt.gz


title: S2 organoids, 90 days, rep2

value: patient


title: S2 iPSC, 8w, rep2

value: 8 weeks


title: S2_corr iPSC, 8w, rep 3


title: S2 iPSC, 8w, rep1


title: S2 organoids, 90 days, rep1


title: H3K27me3 in vehicle-treated control rat rep3

description:
03_Control-3_H3K27me3_rn-dm_i76
HistonePath_rn.txt.gz

value: Sprague-Dawley

value: healthy


title: NR009: Nonresponder_009

value: Lymph node


title: H3K27me3 in vehicle-treated VPA rat rep1

description:
04_Model-Veh-1_H3K27me3_rn-dm_i77
HistonePath_rn.txt.gz

value: prenatal VPA injection

value: vehicle


title: H3K27me3 in vehicle-treated VPA rat rep2

description:
05_Model-Veh-2_H3K27me3_rn-dm_i78
HistonePath_rn.txt.gz


title: H3K27me3 in vehicle-treated control rat rep2

description:
02_Control-2_H3K27me3_rn-dm_i75
HistonePath_rn.txt.gz


title: NR008: Nonresponder_008

value: Nonresponder

value: Brain metastasis


title: S2_corr iPSC, 8w, rep 2


title: NR011: Nonresponder_011

value: Skin metastasis

value: M


title: S2 iPSC, 4w, rep2

value: 4 weeks


title: NR006: Nonresponder_006

value: F

value: Melanoma


title: H3K27me3 in vehicle-treated control rat rep1

description:
01_Control-1_H3K27me3_rn-dm_i74
HistonePath_rn.txt.gz


title: S2_corr iPSC, 8w, rep 1

value: corrected


title: NR005: Nonresponder_005


title: S2_corr iPSC, 4w, rep 3


title: NR010: Nonresponder_010


title: S2 iPSC, 4w, rep1


title: NR004: Nonresponder_004

value: Lung metastasis


title: S2 iPSC, 4w, rep3


title: S2_corr iPSC, 4w, rep 1


title: S1 iPSC, 8w, rep3


title: S1 iPSC, 8w, rep2


title: S2_corr iPSC, 4w, rep 2


title: Myeloid Malignancy Sample N90

city: Baltimore

first: Ying

last: Zou

state: MD

address: 1812 Ashland Ave, Suite 221

department: Department of Pathology

institute: Johns Hopkins University

data_processing:
Binary Cluster files (raw data) from Hiseq or Novarseq instruments are used to generate fastq data using Illumina bcl2fastq software
Genome_build: hg19

source_name: Patient sample

extract_protocol:
DNA was extracted from blood, or bone marrow specimens by conventional methods (Maxwell system; Promega Corporation, Madison, WI)
DNA was captured with Kapa Roche reagents, Integrated DNA Technologies probes,

tag: tissue

value: Bone marrow/Peripheral blood

tag: case

value: 20q deletions


title: Myeloid Malignancy Sample N91


title: S2 iPSC, 8w, rep3

value: patient

value: 8 weeks


title: S2 iPSC, 8w, rep 3


title: HLE-B3_WT_QKI-5_rep2

city: Shanghai

first: Chenxi

last: He

address: 131 Dong'an Rd, Mingdao Bldg, Rm 512

country: China

institute: Fudan University

data_processing:
Basecalls performed using bcl2fastq
ChIP-seq reads (only R1) were aligned to the mm10 genome assembly using bowtie (version 1.2.2)
peaks were called using macs2(v2.1.2) with the following setting: macs2 callpeak -f BAM -g mm -q 0.05 -t ChIP.bam -n NAME -c INPUT.bam
Genome_build: mm10 or hg19
Supplementary_files_format_and_content: bed files were generated using macs2

source_name: HLE-B3 cell line

growth_protocol: HLE-B3 cells were obtained from ATCC (Cat# CRL-11421) and cultured in EMEM containing 20% fetal bovine serum, 50 units/mL penicillin G, and 50 μg/mL streptomycin.

extract_protocol:
Cells were crosslinked, sonicated and  then incubated with Qki-5, Pol II, Srebp2 and normal rabbit IgG antibodies. Protein-DNA complex were washed and ChIP'ed DNA were isolated for library preparation.
Libraries were prepared by KAPA Hyper Prep Kit (KK8504, Kapa Biosystems).

value: Human lens epithelial cells, transformed with an adenovirus 12-SV40 virus hybrid (Ad12-SV40)

tag: passages

value: 20 after received from ATCC

value: Infant less than 1 year

value: Qki-5 (home-made)

tag: genotype

value: wild type


title: S2 iPSC, 8w, rep 2


title: S1 iPSC, 8w, rep1


title: Myeloid Malignancy Sample N88


title: NR007: Nonresponder_007

value: Nonresponder


title: S1 iPSC, 4w, rep3


title: S1 iPSC, 4w, rep2


title: S2 iPSC, 8w, rep 1


title: S1 iPSC, 4w, rep1


title: Myeloid Malignancy Sample N87


title: R011: Responder_011

value: Responder


title: NR001: Nonresponder_001

value: Lymph Node

value: M


title: NR002: Nonresponder_002

value: Anal metastasis

value: F


title: R012: Responder_012

value: Melanoma


title: Myeloid Malignancy Sample N86


title: S2 iPSC, 4w, rep 3

value: 4 weeks


title: NR003: Nonresponder_003

value: Skin metastasis


title: Myeloid Malignancy Sample N89

value: Bone marrow/Peripheral blood

value: 20q deletions


title: S2 iPSC, 4w, rep 1

value: patient


title: H1 hESC, 8w, rep2

value: control

value: 8 weeks


title: H1 hESC, 8w, rep1


title: H1 hESC, 4w, rep3


title: Myeloid Malignancy Sample N85


title: Myeloid Malignancy Sample N82


title: Myeloid Malignancy Sample N83


title: R008: Responder_008

value: Lymph node


title: Myeloid Malignancy Sample N84


title: Myeloid Malignancy Sample N81

value: trisomy / tetrasomy 8


title: R006: Responder_006


title: R007: Responder_007


title: H1 hESC, 4w, rep2


title: H1 hESC, 4w, rep1


title: R010: Responder_010


title: R009: Responder_009


title: S2 iPSC, 4w, rep 2


title: Myeloid Malignancy Sample N80


title: Myeloid Malignancy Sample N79


title: Myeloid Malignancy Sample N77


title: Myeloid Malignancy Sample N78


title: Myeloid Malignancy Sample N76


title: Zcchc8 KO clone 3 rep 3

city: Bethesda

first: Kenneth

last: Yamada

address: BG 30 RM 408 30 CONVENT DR

institute: NIDCR

data_processing:
The samples were analyzed on an Illumina NextSeq500 configured for 40 paired-end reads and the quality of the reads was evaluated using FastQC software.
Read mapping against the recent GENCODE mouse release M24 was performed using the STAR 2.7.3a aligner using the standard mode with mapping parameters derived from the GENCODE project.
Each genomic feature was given a specific identifier in the feature identifier column of the respective new regulatory build gff files (ctcf, enhancer, ocr, promoter, pfr, tfbs).  STAR genome generation was then conducted with the additional parameters --sjdbGTFfeatureExon feature and --sjdbGTFtagExonParentGene ID.
For PROMPTs, the primary assembly gtf file was filtered to remove all feature types except for the “gene” identifier.  This identifier was changed to “prompt” and the genomic coordinates were altered to 3 kb upstream of the start site.  STAR genome generation was then conducted with the additional parameters --sjdbGTFfeatureExon prompt and --sjdbGTFtagExonParentGene ID.
Read counting was performed using the quantMode utility of STAR.
Read counts were filtered to remove low expressing features (genes, PROMPTs, genomic regulatory features, etc., with 5 counts in at least one sample).
Genome_build: GENCODE mouse release M24
Supplementary_files_format_and_content: Tab deliminated txt files with raw counts:
ALL_WT_raw_counts_genes.txt  - raw gene counts including anamolous WT01 sample
raw_counts_genes.txt  - raw gene counts excluding anamolous WT01 sample
raw_counts_ctcf.txt  - raw ctcf counts excluding anamolous WT01 sample
raw_counts_enhancers.txt  - raw enhancer counts excluding anamolous WT01 sample
raw_counts_ocr.txt  - raw open chromatin region counts excluding anamolous WT01 sample
raw_counts_pfr.txt  - raw promoter flanking region counts excluding anamolous WT01 sample
raw_counts_promoters.txt - raw promoter counts excluding anamolous WT01 sample
raw_counts_prompts.txt  - raw PROMPT counts excluding anamolous WT01 sample
raw_counts_tfbs.txt - raw transcription factor binding site counts excluding anamolous WT01 sample

organism: Mus musculus

source_name: SIMS Zcchc8 KO cells

growth_protocol: DMEM + 10% FBS

extract_protocol:
RNA extraction using Trizol and Qiagen Spin columns
RNA integrity was assessed using a Fragment Analyzer (Advanced Analytical) and sequencing libraries were prepared using the Illumina TruSeq Stranded Total RNA library preparation kit according to the manufacturer’s protocols

treatment_protocol: CRISPR/Cas9 KO of either Btbd7 or Zcchc8

value: salivary gland

value: SIMS

value: Zcchc8 KO clone 3


title: R004: Responder_004

value: Responder

value: Ileal metastasis


title: Myeloid Malignancy Sample N75

value: monosomy 5 / 5q deletions

value: 7q deletions


title: R001: Responder_001

value: F

value: Melanoma


title: Zcchc8 KO clone 3 rep 2


title: Zcchc8 KO clone 2 rep 4

value: Zcchc8 KO clone 2


title: Zcchc8 KO clone 3 rep 1


title: CGL

city: Albuquerque

first: Rajanikanth

last: Vadigepalli

state: PA

address: 1 University of New Mexico

institute: University of New Mexico

description:
Neurons from the Left Side of DMV in SHAM surgery animal
BioMark-RTqPCR

data_processing: Samples were normalized through centering aound the median of all genes that passed quality control through melt curve analysis.

label: EvaGreen

source_name: DMV neurons

label_protocol: cDNA labeled with SYBR green according to manufacture's protocol.

extract_protocol: Brainstems were extracted and frozen in OCT, sectioned on a cryostat at 10 um, sections were prepared with rapid IHC for DAPI and NeuN in less than 15 minutes, single neurons were captured using Arcturus CapSure system according to manufacture protocols.  Single cells or cell pools (2-5 single cells) were indiviually lysed and PCR performed according to Fluidigm's Biomark preparation protocols.

treatment_protocol: Male sprague dawley rats were subjected to ligation of the left anterior decending artery after exteriorizing the heart through a small left thoracotomy. Sham surgery included ketamine/xylazine anesthesia and surgical exposure without ligation. n=3-4 biological replicates. All samples were collected after either 1 week or 3 weeks post surgery.

tag: strain

value: Sprague Dawley

tag: animal number

value: CG

tag: side of brain stem

value: L

tag: surgery type

value: SHAM

tag: time point

value: 3 Weeks


title: Myeloid Malignancy Sample N72

value: Bone marrow/Peripheral blood


title: Zcchc8 KO clone 2 rep 2

tag: cell line


title: Myeloid Malignancy Sample N73


title: Zcchc8 KO clone 3 rep 4


title: R002: Responder_002

value: Lung metastasis

value: M

tag: cell type


title: CGK

description:
Neurons from theRight Side of DMV in SHAM surgery animal
BioMark-RTqPCR

value: R


title: R005: Responder_005

value: Adrenal Metastasis


title: Myeloid Malignancy Sample N74

value: trisomy / tetrasomy 8


title: R003: Responder_003

value: Responder

value: Lymph Node


title: CGJ


title: CGF


title: CGI


title: Myeloid Malignancy Sample N71

value: monosomy 5 / 5q deletions

value: 7q deletions


title: CGE


title: Zcchc8 KO clone 2 rep 1

value: salivary gland

value: SIMS


title: Zcchc8 KO clone 1 rep 4

value: Zcchc8 KO clone 1


title: Myeloid Malignancy Sample N70


title: Zcchc8 KO clone 2 rep 3

value: Zcchc8 KO clone 2


title: Zcchc8 KO clone 1 rep 3


title: Zcchc8 KO clone 1 rep 2


title: CGD

value: Sprague Dawley

value: CG

value: L


title: Myeloid Malignancy Sample N68

value: Bone marrow/Peripheral blood


title: Myeloid Malignancy Sample N66

value: 20q deletions


title: Myeloid Malignancy Sample N65


title: Myeloid Malignancy Sample N67


title: Wild type rep 3

source_name: SIMS wild type cells

value: Wild type


title: Wild type rep 4


title: Myeloid Malignancy Sample N64

value: monosomy 7


title: CFL

description:
Neurons from the Left Side of DMV in LAD surgery animal
BioMark-RTqPCR

value: CF

value: LAD

value: 3 Weeks


title: CFK

description:
Neurons from the Right Side of DMV in LAD surgery animal
BioMark-RTqPCR

value: R


title: Myeloid Malignancy Sample N63


title: Wild type rep 2


title: Wild type rep 1


title: CFH


title: CFF


title: Myeloid Malignancy Sample N62

value: monosomy 5 / 5q deletions

value: trisomy / tetrasomy 8


title: Btbd7 KO rep 4

source_name: SIMS Btbd7 KO cells

value: salivary gland

value: Btbd7 KO


title: Zcchc8 KO clone 1 rep 1

value: SIMS

value: Zcchc8 KO clone 1


title: Myeloid Malignancy Sample N69

value: 7q deletions


title: Myeloid Malignancy Sample N61


title: Btbd7 KO rep 3


title: CEI

value: Sprague Dawley

value: CE


title: CFD

value: L


title: CFE


title: CGA

value: CG

value: SHAM


title: Myeloid Malignancy Sample N60

value: Bone marrow/Peripheral blood


title: Btbd7 KO rep 2


title: Btbd7 KO rep 1


title: Myeloid Malignancy Sample N58

value: 20q deletions


title: DNA methylation in TAK-418-treated poly I:C mouse rep2

data_processing:
The reads are mapped to the genome using RRBSMAP using default settings (12bp seed length, max number of mismatches = 0.08 x read length); only unique alignments are kept (if a fragment maps to more than one location, it is discarded)
The methylation ratio values were calculated by the methyratio function in RRBSMAP
Genome_build: rn6 or mm10
Supplementary_files_format_and_content: standard BED file containing methylation ratios of individual CpGs.

extract_protocol:
Brain cortex tissues were removed, and flash frozen on dry ice
Libraries were prepared by Active Motif

tag: disease state

value: prenatal poly I:C injection

tag: treatment


title: CFC

value: CF

value: LAD

value: 3 Weeks


title: CED


title: Myeloid Malignancy Sample H54


title: Myeloid Malignancy Sample H52


title: Myeloid Malignancy Sample H55


title: DNA methylation in vehicle-treated poly I:C mouse rep3

value: vehicle


title: DNA methylation in TAK-418-treated poly I:C mouse rep1


title: CEE

value: R


title: CEF


title: CEG


title: Myeloid Malignancy Sample H51

value: monosomy 5 / 5q deletions


title: CEB


title: DNA methylation in vehicle-treated poly I:C mouse rep2


title: DNA methylation in vehicle-treated poly I:C mouse rep1


title: CEC


title: Myeloid Malignancy Sample N59


title: Myeloid Malignancy Sample H48

value: trisomy / tetrasomy 8


title: Myeloid Malignancy Sample H50


title: DNA methylation in vehicle-treated control mouse rep3

value: healthy


title: DNA methylation in vehicle-treated control mouse rep2


title: CFG


title: CEA


title: DNA methylation in TAK-418-treated poly I:C mouse rep3


title: DNA methylation in vehicle-treated control mouse rep1


title: Myeloid Malignancy Sample H42


title: CDY

value: CD


title: CDX

value: Sprague Dawley


title: Myeloid Malignancy Sample H41


title: CDW


title: CDZ


title: Pooled input of mouse samples

value: pooled

tag: chip antibody


title: H3K9me3 in TAK-418-treated poly I:C mouse rep3

description:
36_Model-CpdH-3_H3K9me3_mm-dm_i69
HistonePath_mm.txt.gz


title: Myeloid Malignancy Sample H40


title: Myeloid Malignancy Sample H38


title: Myeloid Malignancy Sample H36


title: CDV

value: L


title: Myeloid Malignancy Sample H46


title: Myeloid Malignancy Sample H33


title: H3K9me3 in TAK-418-treated poly I:C mouse rep1

description:
34_Model-CpdH-1_H3K9me3_mm-dm_i67
HistonePath_mm.txt.gz


title: H3K9me3 in vehicle-treated poly I:C mouse rep3

description:
33_Model-Veh-3_H3K9me3_mm-dm_i64
HistonePath_mm.txt.gz


title: CDU


title: Myeloid Malignancy Sample H32


title: H3K9me3 in vehicle-treated poly I:C mouse rep2

description:
32_Model-Veh-2_H3K9me3_mm-dm_i61
HistonePath_mm.txt.gz


title: Escherichia coli JW2553 (EF4 mutant)_replicate 2 (RNA-Seq)

city: Beijing

first: Bin

last: Liu

state: Beijing

address: Hai Dian

department: School of Life Sciences

institute: Tsinghua University

description: mRNA

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 BW25113 whole genome using bowtie v2.2.9
Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated using a protocol from Chepelev et al., Nucleic Acids Research, 2009. The number of reads falling in the exons of this meta-transcript were counted and normalized by the size of the meta-transcript and by the size of the library.
Genome_build: BW25113 (Genbank accession: CP009273.1)
Supplementary_files_format_and_content: Excel file includes RPKM values for each Sample

organism: Escherichia coli

source_name: Escherichia coli JW2553 (EF4 mutant)

extract_protocol:
Ribosome footprints were extracted using Rneasy MiNi Kit according to the manual. mRNAs were extracted using TRIZOL.
The libraries were prepared for sequencing using standard protocols

value: 0.2 μg/ml tellurite treatment

value: mutant


title: Myeloid Malignancy Sample H31

value: Bone marrow/Peripheral blood

value: 7q deletions


title: Escherichia coli JW2553 (EF4 mutant)_replicate 1 (RNA-Seq)

description: mRNA


title: H3K9me3 in vehicle-treated poly I:C mouse rep1

description:
31_Model-Veh-1_H3K9me3_mm-dm_i59
HistonePath_mm.txt.gz

value: prenatal poly I:C injection


title: CDL


title: Escherichia coli BW25113_replicate 2 (RNA-Seq)

description: mRNA

source_name: Escherichia coli BW25113

value: wild type


title: CDK

value: LAD


title: Myeloid Malignancy Sample H29


title: Myeloid Malignancy Sample H28

value: 20q deletions


title: CDJ

value: 3 Weeks


title: Escherichia coli BW25113_replicate 1 (RNA-Seq)

description: mRNA


title: Escherichia coli JW2553 (EF4 mutant)_replicate 2 (Ribo-Seq)

description: Ribosome footprints

data_processing:
Library strategy: Ribo-Seq
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 BW25113 whole genome using bowtie v2.2.9
Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated using a protocol from Chepelev et al., Nucleic Acids Research, 2009. The number of reads falling in the exons of this meta-transcript were counted and normalized by the size of the meta-transcript and by the size of the library.
Genome_build: BW25113 (Genbank accession: CP009273.1)
Supplementary_files_format_and_content: Excel file includes RPKM values for each Sample


title: CCJ

value: CC

value: SHAM


title: CCI

description:
Neurons from the Right Side of DMV in SHAM surgery animal
BioMark-RTqPCR

value: R


title: H3K9me3 in vehicle-treated control mouse rep1

description:
28_Control-1_H3K9me3_mm-dm_i55
HistonePath_mm.txt.gz

value: healthy

value: vehicle


title: Myeloid Malignancy Sample H26


title: Escherichia coli JW2553 (EF4 mutant)_replicate 1 (Ribo-Seq)

description: Ribosome footprints


title: CCH

value: Sprague Dawley


title: Myeloid Malignancy Sample H23

value: monosomy 5 / 5q deletions


title: H3K9Ac in TAK-418-treated poly I:C mouse rep3

description:
27_Model-CpdH-3_H3K9Ac_mm-dm_i54
HistonePath_mm.txt.gz


title: CCG


title: Myeloid Malignancy Sample H19

value: monosomy 7


title: H3K9Ac in TAK-418-treated poly I:C mouse rep2

description:
26_Model-CpdH-2_H3K9Ac_mm-dm_i53
HistonePath_mm.txt.gz


title: Escherichia coli BW25113_replicate 1 (Ribo-Seq)

description: Ribosome footprints

value: 0.2 μg/ml tellurite treatment


title: H3K9me3 in vehicle-treated control mouse rep3

description:
30_Control-3_H3K9me3_mm-dm_i57
HistonePath_mm.txt.gz


title: S9

city: Washington

first: Bevan

last: Main

state: District of Columbia

address: 3970 Reservoir Rd NW

department: Neuroscience

institute: Georgetown University

data_processing:
RNA libraries were prepared for sequencing by Psomagen
The raw sequencing reads (FASTQ files) were aligned to the mm10 mouse genome build using the splice-aware read aligner HISAT2.
Transcript is assembled by StringTie with aligned reads based on reference genome model
Expression profiles are represented as read count and normalization value which is based on transcript length and depth of coverage.
The FPKM (Fragments Per Kilobase of transcript per Million Mapped reads) value or the RPKM (Reads Per Kilobase of transcript per Million mapped reads) is used as a normalization value.
Genome_build: mm10
Supplementary_files_format_and_content: Matrix table with contrasts for every gene and every sample

source_name: Saline + Sham_brain

extract_protocol:
Brains were removed, cortex were dissected, flash frozen on dry ice, and RNA was harvested using Trizol reagent.
RNA libraries were prepared for sequencing by Psomagen inc.

tag: age

value: 12 weeks

tag: Sex

value: male

value: Saline + Sham

value: brain; cortex


title: H3K9Ac in TAK-418-treated poly I:C mouse rep1

description:
25_Model-CpdH-1_H3K9Ac_mm-dm_i52
HistonePath_mm.txt.gz

value: prenatal poly I:C injection


title: Myeloid Malignancy Sample H18

value: Bone marrow/Peripheral blood


title: H3K9me3 in vehicle-treated control mouse rep2

description:
29_Control-2_H3K9me3_mm-dm_i56
HistonePath_mm.txt.gz


title: H3K9Ac in vehicle-treated poly I:C mouse rep3

description:
24_Model-Veh-3_H3K9Ac_mm-dm_i50
HistonePath_mm.txt.gz


title: Myeloid Malignancy Sample H17


title: Myeloid Malignancy Sample H14


title: S8


title: H3K9Ac in vehicle-treated poly I:C mouse rep2

description:
23_Model-Veh-2_H3K9Ac_mm-dm_i49
HistonePath_mm.txt.gz


title: S7


title: Myeloid Malignancy Sample H12


title: H3K9Ac in vehicle-treated poly I:C mouse rep1

description:
22_Model-Veh-1_H3K9Ac_mm-dm_i48
HistonePath_mm.txt.gz


title: S6


title: CBL

description:
Neurons from the Left Side of DMV in SHAM surgery animal
BioMark-RTqPCR

value: CB

value: L


title: CCC

value: CC

value: 3 Weeks


title: CCE

value: R

value: SHAM


title: Escherichia coli BW25113_replicate 2 (Ribo-Seq)

description: Ribosome footprints

value: wild type


title: Myeloid Malignancy Sample H11


title: H3K9Ac in vehicle-treated control mouse rep3

description:
21_Control-3_H3K9Ac_mm-dm_i45
HistonePath_mm.txt.gz

value: healthy

value: vehicle


title: S5


title: CBK

value: Sprague Dawley


title: CCF


title: Myeloid Malignancy Sample H9

value: monosomy 5 / 5q deletions


title: H3K9Ac in vehicle-treated control mouse rep2

description:
20_Control-2_H3K9Ac_mm-dm_i44
HistonePath_mm.txt.gz


title: CBJ


title: S4

value: 12 weeks


title: CCD


title: H3K9Ac in vehicle-treated control mouse rep1

description:
19_Control-1_H3K9Ac_mm-dm_i43
HistonePath_mm.txt.gz


title: CBI


title: S3

value: male


title: Myeloid Malignancy Sample H8


title: Myeloid Malignancy Sample H6


title: H3K4me3 in TAK-418-treated poly I:C mouse rep3

description:
18_Model-CpdH-3_H3K4me3_mm-dm_i42
HistonePath_mm.txt.gz


title: S2

value: Saline + Sham


title: S10

value: brain; cortex


title: Myeloid Malignancy Sample H5


title: CBH


title: H3K4me3 in TAK-418-treated poly I:C mouse rep2

description:
17_Model-CpdH-2_H3K4me3_mm-dm_i40
HistonePath_mm.txt.gz

value: prenatal poly I:C injection


title: CAN

description:
Neurons from the Right Side of DMV in LAD surgery animal
BioMark-RTqPCR

value: CA

value: LAD


title: Myeloid Malignancy Sample H1

value: Bone marrow/Peripheral blood


title: H3K4me3 in vehicle-treated poly I:C mouse rep3

description:
15_Model-Veh-3_H3K4me3_mm-dm_i95
HistonePath_mm.txt.gz


title: CBG

description:
Neurons from the Right Side of DMV in SHAM surgery animal
BioMark-RTqPCR

value: CB


title: CAL

value: 3 Weeks


title: CAM

description:
Neurons from the Left Side of DMV in LAD surgery animal
BioMark-RTqPCR

value: L


title: MEM-HFHI-8

source_name: Memantine + HF-HI_brain

value: Memantine + HF-HI


title: MEM-HFHI-9


title: S1


title: H3K4me3 in vehicle-treated poly I:C mouse rep1

description:
13_Model-Veh-1_H3K4me3_mm-dm_i93
HistonePath_mm.txt.gz


title: iFbN_33i_rep3_bulkATACseq

first: Mark

middle: R

last: Cookson

address: 35 Convent Drive

department: National Institute on Aging

institute: National Institutes of Health

description: bulkATACseq_Sample 7

data_processing:
After adapter trimming using NGmerge (v0.3) sequences were aligned to the GRCh38 genome build using bowtie2 (v2.4.1)
Peaks were called using Genrich (v0.6)
Genome_build: hg38
Supplementary_files_format_and_content: Called peaks of signal enrichment in ENCODE narrowPeak format.

source_name: iPSC-derived forebrain neuron cells

extract_protocol:
iFbN were collected at differentiation day 63 with Accutase, then nuclei were extracted from 50,000 cells.
DNA libraries were constructed using custom PCR primers according to the Buenrostro et al. 2015 method.

tag: rs76904798 genotype

value: not available

tag: differentiation protocol

value: iPSC were differentiated to forebrain neuron-like cells according to Burkhardt et al 2013 (PMID 23891805).

value: CS83iCTR-33n1 (33i) iPSC

tag: rna population


title: iFbN_33i_rep1_bulkATACseq

description: bulkATACseq_Sample 5

tag: tissue

value: iPSC were differentiated to forebrain neuron-like cells


title: iFbN_33i_rep2_bulkATACseq

description: bulkATACseq_Sample 6


title: MEM-HFHI-7


title: CAK

value: Sprague Dawley


title: MEM-HFHI-6

value: 12 weeks


title: iMGL_4101_bulkATACseq

description: bulkATACseq_Sample 4

source_name: iPSC-derived microglia cells

extract_protocol:
Live iMicroglia were purified by FACS on differentiation day 28, then nuclei were extracted from 50,000 cells.
DNA libraries were constructed using custom PCR primers according to the Buenrostro et al. 2015 method.

value: CT

value: iPSC were differentiated to microglia-like cells according to Abud et al 2017 (PMID 28426964), McQuade et al 2018 (PMID 30577865).

value: PPMI-4101 iPSC


title: H3K4me3 in vehicle-treated control mouse rep3

description:
12_Control-3_H3K4me3_mm-dm_i92
HistonePath_mm.txt.gz

extract_protocol:
Brain cortex tissues were removed, and flash frozen on dry ice. Tissues were fixed with 1% formaldehyde for 15 min and quenched with 0.125 M glycine.  Lysates were sonicated with a microtip sonicator to shear the DNA. Genomic DNA (Input) was prepared by treating aliquots of chromatin with RNase, proteinase K and heat for de-crosslinking, followed by ethanol precipitation. For each ChIP reaction, 30 ug of chromatin was precleared with protein A agarose beads (Invitrogen). ChIP reaction was set up using precleared chromatin and antibody and incubated overnight at 4 C. Protein A agarose beads were added and incubation at 4 C was continued for another 3 hr. Immune complexes were washed, eluted from the beads with SDS buffer, and subjected to RNase and proteinase K treatment. Crosslinks were reversed by incubation overnight at 65 C, and ChIP DNA was purified by phenol-chloroform extraction and ethanol precipitation.
Illumina sequencing libraries were prepared from the ChIP and Input DNAs by the standard consecutive enzymatic steps of end-polishing, dA-addition, and adaptor ligation. After a final PCR amplification step, the resulting DNA libraries were quantified and sequenced on NextSeq 500.

value: healthy

value: vehicle


title: H3K4me3 in TAK-418-treated poly I:C mouse rep1

description:
16_Model-CpdH-1_H3K4me3_mm-dm_i96
HistonePath_mm.txt.gz

value: prenatal poly I:C injection


title: CAJ

value: CA

value: R

value: LAD


title: iMGL_3453_bulkATACseq

description: bulkATACseq_Sample 3

value: CC

value: iPSC were differentiated to microglia-like cells

value: PPMI-3453 iPSC


title: MEM-HFHI-5

value: male

value: Memantine + HF-HI

value: brain; cortex


title: iMGL_3446_bulkATACseq

description: bulkATACseq_Sample 2

value: PPMI-3446 iPSC


title: H3K4me3 in vehicle-treated control mouse rep1

description:
10_Control-1_H3K4me3_mm-dm_i90
HistonePath_mm.txt.gz


title: CAH

value: Sprague Dawley

value: L

value: 3 Weeks


title: MEM-HFHI-4

value: 12 weeks


title: iMGL_3411_bulkATACseq

description: bulkATACseq_Sample 1

value: iPSC were differentiated to microglia-like cells according to Abud et al 2017 (PMID 28426964), McQuade et al 2018 (PMID 30577865).

value: PPMI-3411 iPSC


title: H3K27me3 in TAK-418-treated poly I:C mouse rep3

description:
09_Model-CpdH-3_H3K27me3_mm-dm_i89
HistonePath_mm.txt.gz

value: prenatal poly I:C injection


title: H3K27me3 in TAK-418-treated poly I:C mouse rep2

description:
08_Model-CpdH-2_H3K27me3_mm-dm_i88
HistonePath_mm.txt.gz


title: MEM-HFHI-3


title: CAG

value: CA


title: MEM-HFHI-2


title: DFL

value: DF

value: 1 Week


title: MEM-HFHI-10


title: H3K4me3 in vehicle-treated control mouse rep2

description:
11_Control-2_H3K4me3_mm-dm_i91
HistonePath_mm.txt.gz

value: healthy

value: vehicle


title: CAI

value: R

value: LAD


title: H3K27me3 in vehicle-treated poly I:C mouse rep3

description:
06_Model-Veh-3_H3K27me3_mm-dm_i86
HistonePath_mm.txt.gz


title: DFK


title: MEM-9

source_name: Memantine+ Sham_brain

value: male

value: Memantine+ Sham


title: H3K27me3 in vehicle-treated poly I:C mouse rep2

description:
05_Model-Veh-2_H3K27me3_mm-dm_i85
HistonePath_mm.txt.gz

source_name: Brain cortex


title: DFJ


title: P61 Blood

city: Paris

first: Paul

last: Gueguen

address: 26 rue D'Ulm

department: Inserm U932

country: France

institute: Institut Curie

data_processing:
sample de-multiplexing and alignment was performed using Cell Ranger Single Cell Software Suite (v2.0.1) and with for and (v3.0.2) for P57, P58, P60 and P61.
Genome_build: GRCh38
Supplementary_files_format_and_content: Processed files are filtered contigs annotations files which are given by Cellranger pipeline.

source_name: Blood sample

extract_protocol:
TCR libraries from P47, P55, P57, P58 and P61 was extracted according to the Single Cell VDJ 5' Reagent Kits
TCR enrichment from amplified cDNA
5' for paired single cell TCR-seq

treatment_protocol: Tumor and adjacent normal lung tissue samples were gently cut in approximately 1mm3 pieces. Tissues were digested enzymatically, by an incubation of 20-40 minutes, based on the size of the tissue, at 37oC in agitation, in C02-independent medium (Invitrogen) with Collagenase I (2mg/ml, Sigma-Aldrich), Hyaluronidase (2mg/ml, Sigma-Aldrich) and DNase (25μg/ml, Sigma-Aldrich). The tissue pieces were gently triturated with a 20 ml syringe plunger on a 40 μ m cell strainer (BD) in 1X PBS (Invitrogen) with 1% fetal bovine serum (FBS) and 2mM EDTA (Gibco) until uniform cell suspensions were obtained. The suspended cells were subsequently centrifuged for 10 minutes at 400g and resuspended in PBS 1X buffer with 1% FBS and 2mM EDTA (Gibco). Isolation of tumor infiltrating lymphocytes and peripheral blood mononuclear cells was performed using Ficoll-paque PLUS solution (Sigma-Aldrich). CD3+ T cell isolationwas performed using CD3 microbeads and MACS separation (Miltenyi). Susequently dead cells and debris was removed following SOP (Dead cell removal kit; Debris removal kit; Miltenyi Biotec). Purified CD3+ T cells were resuspended in 1X PBS buffer with 0.04% BSA.

tag: tumor stage

value: Blood sample

tag: smoking status

tag: tumor type

value: M


title: H3K27me3 in vehicle-treated poly I:C mouse rep1

description:
04_Model-Veh-1_H3K27me3_mm-dm_i84
HistonePath_mm.txt.gz

data_processing:
Sequences: Standard Illumina software base-calling and quality-control filtering was applied
Sequences (75 nt reads, single end) were aligned to the mouse genome (mm10) or rat genome (rn5) using the BWA algorithm (v0.7.12, default parameters).
Aligns were extended in silico at their 3’-ends to a length of 200 bp, which is the average genomic fragment length in the size-selected library, and assigned to 32-nt bins along the genome. The resulting histograms (genomic “signal maps”) were stored in bigWIG files.
Peak locations were determined using the MACS algorithm (v2.1.0) with a cutoff of pvalue = 1E-7.
Signal maps and peak locations were used as input data to Active Motifs proprietary analysis program, which creates excel tables containing detailed information on sample comparison, peak metrics, peak locations and gene annotations
Genome_build: rn5 or mm10
Supplementary_files_format_and_content: Peak metrics, peak locations and gene annotations in text format

tag: strain


title: DFH

value: Sprague Dawley

tag: animal number

tag: side of brain stem

value: L

tag: surgery type


title: P61 Juxta

source_name: NSCLC juxta tissue

value: NSCLC juxta tissue


title: MEM-7

value: 12 weeks

value: brain; cortex


title: DFG

tag: time point


title: H3K27me3 in TAK-418-treated poly I:C mouse rep1

description:
07_Model-CpdH-1_H3K27me3_mm-dm_i87
HistonePath_mm.txt.gz

tag: disease state

value: prenatal poly I:C injection


title: MEM-6

tag: treatment


title: MEM-HFHI-1

value: Memantine + HF-HI


title: DFE

value: DF


title: P60 Blood

value: smoker

value: ADK


title: H3K27me3 in vehicle-treated control mouse rep1

description:
01_Control-1_H3K27me3_mm-dm_i80
HistonePath_mm.txt.gz

value: healthy

value: vehicle

tag: chip antibody


title: H3K27me3 in vehicle-treated control mouse rep2

description:
02_Control-2_H3K27me3_mm-dm_i81
HistonePath_mm.txt.gz


title: P61 Tumor

source_name: NSCLC tumor tissue

value: NSCLC tumor tissue


title: MEM-8

tag: age

value: male


title: P60 Tumor

tag: Sex


title: MEM-5

value: Memantine+ Sham


title: DFI

description:
Neurons from the Right Side of DMV in LAD surgery animal
BioMark-RTqPCR

value: R

value: LAD

value: 1 Week


title: DFD

value: Sprague Dawley

value: L


title: P60 Juxta

value: NSCLC juxta tissue

value: M


title: P61_Blood

description: 5' RNA

data_processing:
sample de-multiplexing and alignment was performed using Cell Ranger Single Cell Software Suite (v2.0.1) and with for and (v3.0.2) for P57, P58, P60 and P61.
EmptyDrops R package (Lun et al. 2019) was used to call droplets for the samples processed without Cellranger v2.0.1
Genome_build: GRCh38
Supplementary_files_format_and_content: Processed files are raw count UMI matrices which are given by Cellranger pipeline and integrated Seurat objects with the original uncorrected matrix, in the "RNA" assay, and the batch-corrected matrix in the "integrated" assay

extract_protocol:
RNA from P34, P35, P42, P43 and P46 was extracted according to the Single Cell 3' Reagent Kits V2. RNA from P47, P55, P57, P58 and P61 was extracted according to the Single Cell VDJ 5' Reagent Kits
3' single cell RNA-seq; 5' for paired single cell RNA/TCR-seq

value: Blood sample


title: H3K4me3 in vehicle-treated poly I:C mouse rep2

description:
14_Model-Veh-2_H3K4me3_mm-dm_i94
HistonePath_mm.txt.gz

value: prenatal poly I:C injection


title: MEM-3

value: 12 weeks

value: brain; cortex


title: mESCs, RNASeq_Rep3

city: Philadelphia

first: Phillip

last: Wulfridge

state: Pennsylvania

address: 3601 Spruce St

institute: The Wistar Institute

data_processing:
Alignment (MapR, BisMapR): Bowtie2
Reads from .bam files were strand-separated using the calls "samtools view | awk '$1~"@" || $2 ~/^(99|147)$/{print}'" for forward strand and "samtools view | awk '$1~"@" || $2 ~/^(83|163)$/{print}'" for reverse strand.
Alignment (RNA-Seq): STAR
RNA expected counts were generated with RSEM.
Bigwig files were made from composite and strand-separated .bam files using "deepTools bamCoverage --extendReads --binSize 5", an ENCODE blacklist file for biased regions (Amemiya et al. Nature 2019), and a scale factor based on the total number of paired reads in the composite.
Genome_build: mm10
Supplementary_files_format_and_content: bigwig files compatible with UCSC genome browser and IGV, TPM output in .counts tab-delimited for RNA-Seq alignment

growth_protocol: Mouse embryonic stem cells were grown in DMEM medium with 15% FBS, 25mM HEPES, 1x GlutaMAX, 1x non-essential amino acids, 100 U/mL penicillin-steptomycin, LIF, 3µM GSK inhibitor, 1µM MEK1/2 inhibitor at 37 ˚C.

extract_protocol:
R-loops were collected from supernatant after treatment with MapR protocol (Yan et al. Cell Reports 2019).
Total RNA was collected using Trizol (Invitrogen) and digested with Turbo Dnase (Ambion). rRNA was selectively depleted using FastSelect (Qiagen).
rRNA-depleted RNA was converted to cDNA using the Ultra II Directional RNA kit for Illumina (NEB #E7760).
BisMapR converted R-loops was converted to dsDNA using the Second Synthesis buffer and enzyme mix from Ultra II Directional RNA kit for Illumina (NEB #E7760).
DNA samples were end-repaired using End-Repair Mix (Enzymatics), A-tailed using Klenow exonuclease minus (Enzymatics), purified with MinElute columns (Qiagen), and ligated to Illumina adapters (NEB E7600) with T4 DNA ligase (Enzymatics).  Size selection for fragments >150 bp was performed with AMPure XP (Beckman Coulter).  Libraries were PCR amplified with dual index barcode primers for Illumina sequencing (NEB E7600) using Q5 DNA polymerase (NEB M0491) and purified with MinElute.

treatment_protocol: For BisMapR treatment, R-loops were incubated with 130µL CT Conversion Reagent from the EZ DNA Methylation-Gold Kit (Zymo) at 25 ˚C for 3 hours, then column purified with EZ DNA Methylation-Gold Kit.

tag: cell line

value: ES cell line E14

tag: protocol

value: RNA-Seq


title: H3K9me3 in TAK-418-treated poly I:C mouse rep2

description:
35_Model-CpdH-2_H3K9me3_mm-dm_i68
HistonePath_mm.txt.gz


title: mESCs, RNASeq_Rep2


title: MEM-2


title: P61_Juxta

description: 5' RNA


title: mESCs, RNASeq_Rep1


title: DFA

value: DF


title: DEL

description:
Neurons from the Left Side of DMV in SHAM surgery animal
BioMark-RTqPCR

value: DE

value: SHAM


title: MEM-1

value: male

value: Memantine+ Sham


title: DEK

description:
Neurons from the Right Side of DMV in SHAM surgery animal
BioMark-RTqPCR

value: R


title: MEM-10


title: P60_Blood

description: 5' RNA

value: smoker

value: ADK


title: MEM-4


title: HFHI-9

source_name: Saline + HF-HI_brain

value: Saline + HF-HI


title: P60_Juxta

description: 5' RNA

value: NSCLC juxta tissue


title: mESCs, BisMapR_RNaseA

value: BisMapR

tag: fusion protein

value: GST-RHΔ-Mnase


title: DEJ

value: Sprague Dawley

value: L

value: 1 Week


title: CDS

value: CD

value: LAD

value: 3 Weeks


title: DFF

description:
Neurons from the Left Side of DMV in LAD surgery animal
BioMark-RTqPCR

value: DF


title: mESCs, MapR_Rep2

value: ES cell line E14

value: MapR


title: HFHI-8

value: 12 weeks

value: male

tag: tissue

value: brain; cortex


title: P60_Tumor

description: 5' RNA

tag: tumor stage

value: NSCLC tumor tissue

value: smoker

value: ADK

value: M


title: HFHI-7

value: Saline + HF-HI


title: DEH

value: Sprague Dawley

value: DE

value: SHAM


title: P58_Blood

description: 5' RNA

value: Blood sample

tag: smoking status

value: F


title: mESCs, BisMapR_Rep2

value: BisMapR

value: GST-RHΔ-Mnase


title: P61_Tumor

description: 5' RNA

tag: tumor type


title: DEI

tag: strain

value: R

value: 1 Week


title: DEG

tag: animal number


title: HFHI-5

value: 12 weeks


title: DFB

value: DF

tag: side of brain stem

value: L

value: LAD


title: P58_Tumor

description: 5' RNA

value: NSCLC tumor tissue

value: smoker


title: mESCs, BisMapR_Rep1

value: ES cell line E14


title: H3K27me3 in vehicle-treated control mouse rep3

description:
03_Control-3_H3K27me3_mm-dm_i82
HistonePath_mm.txt.gz

value: healthy

value: vehicle


title: HFHI-4

value: male

value: Saline + HF-HI

value: brain; cortex


title: mreCD 3

city: Jonesboro

first: Robert

middle: Colquhoun

last: Shields

state: Arkansas

address: Lab Science East

department: Biological Sciences

institute: Arkansas State University

description:
repeat 3 of 3
6-mreCD_S21

data_processing:
Step 1: FASTQ Groomer
Step 2: FASTQ Quality Trimmer
Step 3: Cutadapt removing GATCGGAAGAGCACACGTCTGAACTCCAGTCAC
Step 4: Map with Bowtie for Illumina
Step 5: sort
Step 6: htseq-count (see our publication for details)
Genome_build: NC_004350.2
Supplementary_files_format_and_content: Counts obtained from step 6: htseq-count, listed along side each open reading frame

organism: Streptococcus mutans

source_name: bacterial cells_ΔmreCD

growth_protocol: Bacterial cells used for extraction of RNA were grown in BHI medium, in the presence of 5% CO2 at 37°C without agitation, until they reached mid-exponential phase (OD600 ≈ 0.5)

extract_protocol:
Cells were harvested by centrifugation at 4°C for 5 min, treated with bacterial RNAprotect Bacteria Reagent (Qiagen), and immediately stored at -80°C. Total RNA was extracted from bacterial cells using the RNeasy Mini kit (Qiagen), with chromosomal DNA removed using a DNase I kit (Qiagen). To remove 16S and 23S rRNAs, 10 µg of total RNA was treated using the MICROBExpressTM Bacterial mRNA Enrichment Kit
cDNA libraries were created from 100 ng enriched mRNA samples using the NEBNext Ultra Directional RNA Library Prep kit for Illumina and NEBNext multiplex Oligos for Illumina (New England Biolabs, Ipswich, MA) following instructions from the supplier.
RNA-Seq: single-ended short reads were obtained from a set of bar-coded cDNA libraries

tag: strain background

tag: genotype/variation

value: lacking the mreCD gene

tag: media

value: Brain Heart Infusion

tag: growth point


title: HFHI-2

tag: treatment


title: P57_Tumor

description: 5' RNA

value: pT3 N0

value: ADK

value: M


title: HFHI-3

tag: age

value: 12 weeks


title: P55_Tumor

description: 5' RNA

value: pT2b N0

value: CE

tag: Sex


title: DEE

value: Sprague Dawley

value: DE

value: R

tag: surgery type

value: SHAM

tag: time point

value: 1 Week


title: mreCD 1

description:
repeat 1 of 3
4-mreCD_S7


title: DEB

value: L


title: DEC


title: mESCs, MapR_Rep1

tag: cell line

value: ES cell line E14

tag: protocol

value: MapR

tag: fusion protein

value: GST-RHΔ-Mnase


title: HFHI-6

value: male

value: Saline + HF-HI

tag: tissue

value: brain; cortex


title: mreCD 2

description:
repeat 2 of 3
5-mreCD_S24

value: lacking the mreCD gene

value: Brain Heart Infusion


title: UA159 3

description:
repeat 3 of 3
3-UA159_S3

source_name: bacterial cells_WT

value: wild type


title: Air-liquid interface grown bronchioalveolar cells

city: Utrecht

first: Mauro

last: Muraro

state: Utrecht

address: Uppsalalaan, 8

country: Netherlands

institute: Single Cell Discoveries

data_processing: BCL files resulting from sequencing were transformed to FASTQ files with 10x Genomics Cell Ranger mkfastq. FASTQ files were mapped with Cell Ranger count. During sequencing, Read 1 was assigned 150 base pairs, but only the first 28 bp were used for identification of the Illumina library barcode, cell barcode and UMI. R2 was used to map to a custom made reference transcriptome of Hg38 with additional sequences for SARS-CoV-2 added. Filtering of empty barcodes was done using Cell Ranger. Data filtering, normalization and clustering was performed using the Seurat workflow (Seurat v3.2.2).

source_name: Bronchioalveolar cells

growth_protocol: Bronchioalveolar cultures were grown from human lung bud tip organoids and differentiated  at air-liquid interface in transwell inserts in the presence of canalicular stage fetal lung mesenchymal cells  in the basal compartment

extract_protocol:
Cultures were incubated with TrypLE express for 20 min at 37 C 5% CO2 and cells were dissociated by pipetting. Cells were washes once in cold PBS and resuspended in 200  L ice-cold PBS. Ice-cold methanol was added dropwise to the cell pellet for fixation. Cells were stored at -20 C until further processing. Single-cell mRNA sequencing was performed according to standard 10x Genomics 3  V3.1 chemistry protocol. Prior to loading the cells on the 10x Chromium controller, cells were rehydrated in rehydration buffer and counted to assess cell concentration. Cells were loaded and the resulting sequencing libraries were prepared following standard 10x Genomics protocol. The DNA libraries was paired-end sequenced on an Illumina Novaseq S4, with a 2x150 bp Illumina kit.
10x Genomics 3' transcriptome

treatment_protocol: Cells were untreated

tag: cell type

value: Lung bud tip organoid derived bronchioalveolar cells

value: Lung

value: untreated


title: UA159 2

description:
repeat 2 of 3
2-UA159_S8


title: HFHI-10

tag: strain

value: 12 weeks


title: HFHI-1


title: DEF

source_name: DMV neurons

value: Sprague Dawley

tag: animal number

value: DE


title: DEA

tag: side of brain stem

value: R

value: SHAM

value: 1 Week


title: NLPC_WT_Qki-5_rep2

growth_protocol: Neural stem cell-derived lens progenitor-like cells (NLPCs) were differentiated from neural stem cells in DMEM/F-12 supplemented with 0.05% BSA, 1% nonessential amino acids, 50 units/mL penicillin G, and 50 μg/mL streptomycin, 1X N2 and 1X B27 with two-step growth factor treatment, 1st step with 100 ng/ml Noggin and 2nd step with 100 ng/ml bFGF, 20 ng/ml BMP4, and 20 ng/ml BMP7.

value: Neural stem cell-derived lens progenitor-like cells (NLPCs) differentiated from neural stem cells

tag: passages

value: FvB/C57BL6 hybrid

tag: chip antibody

value: Qki-5 (home-made)

tag: genotype


title: DDL

value: DD

value: L


title: DED


title: retina, PKD + Clodronate (rep4)

city: Mannheim

first: Matthias

last: Kolibabka

state: Baden-Wurttemberg

address: Theodor-Kutzer-Ufer 1-3

department: 5th Medical Department

institute: University Hospital Mannheim

data_processing:
Threshold quantification cycles (Cq) were obtained using the manufacturer's Viia7 software and analyzed in the ArrayStudio software package (Version 9, Omicsoft Corporation).
Gene expression levels were obtained by subtracting the average normalization value of four reference genes (18S, B2m, Mrlp19 and Ywhaz) from the individual Cq values.
Fold change 2^-ΔΔCq, where -ΔΔCq = -[ΔCqtest -MV(ΔCqReferenceGenes)]
Samples where transcripts did not reach the threshold within 35 cycles were excluded from the calculation for this particular gene.
Fold Change worksheet reports PKD/SD and PKD + Clodronate/PKD ratios (Reference genes excluded).

label: FAM

source_name: retina, clodronate

label_protocol: cDNA was generated using a high capacity RNA-to-cDNA Kit according to manufacturer's protocol. A quantitative pre-amplification step of 14 cycles was performed using a Taqman Preamp Master Mix Kit according to the manufacturer's protocol. Real-time PCR was performed in parallel using TaqMan microfluidic card technology in a Viia7 thermocycler with a maximum of 40 cycles.

growth_protocol: At 4 and 8 weeks of age, Sprague Dawley and PKD2(HA-1/703-CMV-247) rats were injected with 3 µl solution according to the treatment groups. Uninjected animals served as control groups. Animals were housed with a 12-/12-h light/dark cycle with free access to food and water. Termination occured at 9 weeks of age, 1 week after the second injection.

extract_protocol: Trizol extraction of total RNA was performed according to the manufacturer's instructions.

treatment_protocol: Enucleated Eyes were snapp frozen in liquid nitrogen and stored at -80°C till further preperation. Extraction of the retina was performed at 0°C and immediately suspended in Trizol.

value: Clodronate


title: retina, PKD + Clodronate (rep5)


title: DDK


title: NLPC_WT_IgGR

source_name: Neural stem cell-derived lens progenitor-like cells (NLPCs)

value: normal rabbit IgG (2729, CST)


title: Male Cocaine Rep 2

city: Greenwood

first: Vijay

last: Shankar

state: South Carolina

address: 115 Milford Pines Drive

institute: Clemson University

data_processing:
The mkfastq pipeline within Cell Ranger v3.1 (10X Genomics, Pleasanton, CA) was used to convert BCL files from the sequence run folder to demultiplexed FASTQ files. Release 6 version of the Drosophila melanogaster reference GCA_000001215.4 from NCBI Genbank was indexed using the mkref pipeline and used for alignment using the count pipeline within Cell Ranger v3.1 with the expected cell count parameter set to 5,000 cells.
Raw expression counts output for each sample from the Cell Ranger pipeline was imported and analyzed using the Seurat v3 package in R. Each sample was filtered to remove genes that are expressed in less than 5 cells and to remove cells with fewer than 300 or greater than 2,500 RNA features. Counts were normalized by regularized negative binomial regression using the scTransform pipeline.
Integration of all samples was performed using the SCT method. RunUMAP and FindNeighbors functions were used with 10 dimensions to ordinate expression space and reduce data dimensionality. To identify the cell-type clusters within the dataset, unsupervised clustering using the FindClusters function and a resolution of 0.8 was used. The resolution value was selected by iterating from 0.4 to 2.0 and identifying the value at which there is significant change in the granularity of cluster assignment. In order to identify marker genes for each cluster, DefaultAssay was set to “RNA”, values were normalized using the NormalizeData function, the function FindAllMarkers run on the integrated set with a logFC threshold of 0.5 (default). The top three genes with positive expression for each cluster were extracted and used for cell-type characterization.
Genome_build: GCA_000001215.4 R6.13
Supplementary_files_format_and_content: UMI barcodes, feature set and counts matrix

organism: Drosophila melanogaster

source_name: Drosophila brains

extract_protocol:
Brains were dissected from each fly immediately after it consumed the designated amount of sucrose or cocaine solution. We used a dissociation protocol modified from Croset et al. (2018) and Davie et al. (2018). We dissected brains in cold D-PBS (Gibco, Thermo Fisher Scientific, Waltham, MA) and collected them into 1.7ml tubes in cold Schneider’s medium (Gibco). We collected 20 brains per sample within one hour. We collected eight samples of 20 brains from males and females exposed to cocaine or sucrose, with two biological replicates per treatment and sex. We replaced the D-PBS in the dissection dish after dissecting 2 brains to ensure that it stayed cold and we used separate drops of buffer for decapitation and brain dissection to avoid contaminating the brain samples. We centrifuged the samplesn at 300xg at 4oC for 5 min and removed the supernatant. We then added 450µl of collagenase solution (50 ul of fresh 25mg/ml collagenase (Gibco) in sterile water + 400µl of Schneider’s medium), flicked the tube gently and allowed the brains to incubate at room temperature for 30 min. We replaced the collagenase solution after centrifugation with PBS + 0.04% BSA (NEB, Ipswich, MA). We mechanically dissociated the brains slowly and gently, using stepwise trituration - P200 pipette 5 times, 23G needle pre-wetted with PBS + BSA 5 times, and 27G pre-wetted needle 5 times. We passed the suspension through a pre-wetted 10µm strainer (Celltrics, Görlitz, Germany) aided by gentle tapping. We added 50ul of PBS+BSA to aid flow of the suspension through the strainer. We counted live cells using a hemocytometer with trypan blue exclusion. We proceeded with GEM generation using the Chromium controller (10X Genomics, Pleasanton, CA) if we had a live cell count of > 500 live cells/µl.
We made libraries after GEM generation in accordance with 10X Genomics v3.1 protocols. We determined fragment sizes using Agilent Tapestation kits (Agilent, Santa Clara, CA) - d5000 for amplified cDNA and d1000 for libraries. However, the samples were not diluted at either step since these were not high concentration libraries. We measured the concentrations of amplified cDNA and the final libraries using a Qubit 1X dsDNA HS kit, also without dilution. In addition to Qubit, we quantified the final library concentrations using a qPCR based library quantification kit (KAPA Biosystems, Roche, Basel, Switzerland) in order to measure the concentration of fragment sizes of interest in accordance with the manufacturer’s recommendations. We used 12 cycles for the cDNA amplification and 14 cycles for indexing PCR. We sequenced the final libraries on an S1 flow cell using a Novaseq (Illumina, Inc., San Diego, CA) according to the manufacturer’s instructions.

treatment_protocol: We collected 200 Canton S (B) flies between 3 and 5 days old using CO2 anesthesia, sexes separately. We placed them individually in culture vials containing cornmeal/yeast/molasses/agar culture medium (Genesee Scientific, Inc., San Diego, CA) and allowed them to recover for 24 hours before experimentation. Between 3:00-5:00 PM on the day before the assay, we transferred the flies to vials containing 1.5% agar (Sigma Aldrich, St. Louis, MO) in which a capillary (VWR International, Radnor, PA: 12.7 cm long, 5 μl total volume) filled completely with a solution of 4% sucrose (Sigma Aldrich) and 1% yeast (Fisher Scientific, Hampton, NH) was inserted. The next morning, we replaced the sucrose capillaries for 100 flies with capillaries containing 4% sucrose supplemented with 1 µg/µL of cocaine and 1% yeast; for the other 100 flies, we replaced the sucrose capillaries with fresh 4% sucrose and 1% yeast with no drug. A droplet of mineral oil (Sigma Aldrich) was added to the top of each capillary to minimize evaporation. We collected the first 40 flies that consumed 0.53 µL of cocaine and the first 40 flies that consumed 0.53 µL of sucrose, corresponding to an 8 mm reduction in the height of the solution in the capillary. All experiments were carried out between 8 AM and 11 AM. Flies were allowed to feed for no more than 2 hours.

value: Brains

value: Male

value: Cocaine

value: CantonSB


title: DDJ


title: 43°C treat-3 [H-3_miRNA]

city: Bengbu

first: Meng

last: Liang

state: Anhui

address: Donghai road 2600

institute: bengbu Medical College

data_processing:
Clean reads were obtained by filtering original reads with Solexa CHASTITY quality control. Clean reads were removed adaptor by Cutadapt and leave a length of ≥15 nt tag to get trimmed reads.
Known miRNA was quantified for all trimmed reads using miRDeep2 software. Based on CPM standardized miRNAs, R software edgeR was used for differential expression calculation and differential miRNAs screening.
Genome_build: mm9
Supplementary_files_format_and_content: Excel files include FPKM values for each Sample.

source_name: 43°C treat_testis

extract_protocol:
Total RNA was extracted from testis of heat exposure using Trizol reagent.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: The scrotum were placed in water bath for 25 minutes at 33 °C in the control group or for 25 minutes at 43 °C in the high temperature group.

value: ICR

value: 43 °C treat

value: Testis


title: retina, PKD + Clodronate (rep3)


title: retina, PKD + Clodronate (rep2)


title: Male Cocaine Rep 1


title: 43°C treat-2 [H-2_miRNA]


title: Female Cocaine Rep 2

value: Female


title: 33°C treat -3 [NC-3_miRNA]

source_name: 33°C treat_testis

value: 33 °C treat


title: retina, PKD + Clodronate (rep1)


title: UA159 1

description:
repeat 1 of 3
1-UA159_S15

tag: strain background

tag: genotype/variation

value: wild type

tag: media

value: Brain Heart Infusion

tag: growth point


title: retina, PKD (rep5)

source_name: retina, control

tag: treatment

value: Control


title: DDH

value: Sprague Dawley

tag: surgery type

value: SHAM

tag: time point

value: 1 Week


title: Male Sucrose Rep 1

tag: Sex

value: Sucrose


title: Female Sucrose Rep 2


title: retina, PKD (rep3)


title: retina, PKD (rep6)


title: Female Cocaine Rep 1


title: 43°C treat-1 [H-1_miRNA]


title: Male Sucrose Rep 2


title: DDG

value: DD

value: R


title: retina, PKD (rep4)


title: retina, PKD (rep2)


title: DDC


title: 33°C treat -2 [NC-2_miRNA]


title: 43°C treat-2

data_processing:
Solexa Pipeline Version 1.8 (off-line Base Caller Software, Version 1.8) was used for image processing and base recognition. FastQC software evaluated sequencing quality of reads after adaptor removal (Cutadapt was used to remove 3 'and 5' adaptors). Hisat2 software was used for comparison to the reference genome.
Transcriptional abundance was assessed using StringTie software with reference to official database annotation information.R software Ballgown was used to calculate the FPKM of gene expression level and then screen out the differentially expressed genes.
Genome_build: mm9
Supplementary_files_format_and_content: Excel files include FPKM values for each Sample.


title: 33°C treat -1 [NC-1_miRNA]


title: DDB

value: L


title: DDD


title: DDE


title: Female Sucrose Rep 1

value: Brains


title: DCP

value: DC

value: LAD


title: DDF


title: HLE-B3_QKI KO_Input

data_processing:
Basecalls performed using bcl2fastq
ChIP-seq reads (only R1) were aligned to the hg19 genome assembly using bowtie (version 1.2.2)
peaks were called using macs2(v2.1.2) with the following setting: macs2 callpeak -f BAM -g hs -q 0.05 -t ChIP.bam -n NAME -c INPUT.bam
Genome_build: hg19
Supplementary_files_format_and_content: bed files were generated using macs2

source_name: HLE-B3 cell line

extract_protocol:
Cells were crosslinked, sonicated and  then incubated with Qki-5, Pol II, and Srebp2 antibodies. Protein-DNA complex were washed and ChIP'ed DNA were isolated for library preparation.
Libraries were prepared by KAPA Hyper Prep Kit (KK8504, Kapa Biosystems).

tag: cell line

value: Human lens epithelial cells, transformed with an adenovirus 12-SV40 virus hybrid (Ad12-SV40)

value: 20 after received from ATCC

tag: age

value: Infant less than 1 year

value: QKI KO


title: HLE-B3_QKI KO_POL II

tag: cell type

value: RNA polymerase II CTD repeat YSPTSPS (Abcam, Cat# ab817)


title: 43°C treat-3

tag: strain

value: ICR

value: 43 °C treat

tag: tissue

value: Testis


title: HLE-B3_WT_Input

value: wild type


title: DCO

data_processing: Samples were normalized through centering aound the median of all genes that passed quality control through melt curve analysis.

value: Sprague Dawley

tag: animal number

tag: side of brain stem

value: 1 Week


title: 43°C treat-1


title: 33°C treat -3

value: 33 °C treat


title: HLE-B3_WT_POL II

tag: passages

tag: chip antibody


title: retina, PKD + Controlliposomes (rep6)

source_name: retina, controlliposomes

value: Controlliposomes


title: DCM

value: R


title: retina, PKD (rep1)

value: Control


title: HLE-B3_QKI KO_SREBP2

value: Srebp2 (10007663, Cayman Chemical)

tag: genotype


title: retina, PKD + Controlliposomes (rep3)


title: retina, PKD + Controlliposomes (rep4)


title: 33°C treat -2


title: retina, PKD + Controlliposomes (rep5)


title: DCN

value: DC

value: L

value: LAD


title: HLE-B3_WT_QKI-5

value: Human lens epithelial cells, transformed with an adenovirus 12-SV40 virus hybrid (Ad12-SV40)

value: 20 after received from ATCC

value: Infant less than 1 year

value: Qki-5 (home-made)


title: 33°C treat -1

value: ICR


title: 661W_TRPM1

city: Tainan

first: Che-Hung

last: Shen

state: Taiwan

address: No. 367, Sheng-Li Rd.,, North Dist.,

department: National Institute of Cancer Research

country: Taiwan

institute: National Health Research Institutes

description: 661W cells stably expressing 3xFlag-TRPM1

data_processing:
Clean reads were aligned to the mouse genome (GRCm38) using HISAT2 (version 2.1.0) after removing low-quality reads.
The differential expression of genes between TRPM1-overexpression and control cells was computed using the fragments per kilobase of transcript per million mapped reads calculated by featureCounts (version 2.0.0). Raw read counts were imported into edgeR (version 3.28.1) and analyzed by using R package of DESeq (version 1.40.0).
Genes with false discovery rate (FDR) p-value < 0.05 adjusted by using Benjamini–Hochberg (BH) method were considered as differentially expressed genes (DEGs).
Gene set enrichment analysis of the genes differentially expressed upon TRPM1 expression was done using the Gene Set Knowledgebase（GSKB）hallmark gene sets.
Genome_build: GRCm38
Supplementary_files_format_and_content: DEG_result_table in excel file format contains ensembl_gene_id, log2FoldChange, pvalue, padj, length, symbol, entrezgene, description, norm.count.mean, and baseMean for every gene and every sample
Supplementary_files_format_and_content: GSEA_GO_result_table in excel file format contains ID, Description, setSize, enrichmentScore, NES, pvalue, p.adjust, qvalues, rank, leading_edge, core_enrichment

growth_protocol: Cells were were maintained in DMEM (HyClone) containing 10% fetal bovine serum (FBS; Hyclone) and 100U/ml penicillin/streptomycin (Gibco). Cells were tested negatively for mycoplasma with the MycoSensor PCR Assay Kit (Agilent Technologies).

extract_protocol:
RNAs were isolatedusing the RNeasy Mini kit (Qiagen).
RNA-seq libraries were prepared using the KAPA mRNA HyperPrep Kit (KAPA Biosystems, Roche, Basel, Switzerland) and validated using the Qsep 100 DNA/RNA Analyzer (BiOptic Inc., Taiwan). Libraries were sequenced on a NovaSeq 6000 sequencer (Illumina, CA, USA)

treatment_protocol: Normal cell culture condition

value: photoreceptor cell

tag: genotype/variation


title: HLE-B3_WT_SREBP2

value: wild type


title: 661W_Vec

description: 661W cells stably expressing control vector

value: control vector


title: DCT

extract_protocol: Brainstems were extracted and frozen in OCT, sectioned on a cryostat at 10 um, sections were prepared with rapid IHC for DAPI and NeuN in less than 15 minutes, single neurons were captured using Arcturus CapSure system according to manufacture protocols.  Single cells or cell pools (2-5 single cells) were indiviually lysed and PCR performed according to Fluidigm's Biomark preparation protocols.

value: Sprague Dawley

value: 1 Week


title: retina, PKD + Controlliposomes (rep1)

value: Controlliposomes


title: DCR


title: retina, PKD + Controlliposomes (rep2)


title: retina, PKD + PBS (rep5)

source_name: retina, PBS

value: PBS


title: In-cell M2-seq

city: Stanford

first: Gun Woo

last: Byeon

state: CA

address: 279 Campus Dr., Beckman Center B309

institute: Stanford University

description: cDNA amplicon sequencing

data_processing:
After bcl conversion and demultiplexing with Illumina bcl2fastq, adaptor sequences are further trimmed using cutadapt. Overlapping paired-end reads are merged using BBMerge. Trimmed and merged reads are aligned to indexed reference of amplicon sequences using Bowtie2 with the following parameters: -D 20 -R 3 -N 0 -L 20 -i S,1,0.50 --mp 3,1 --rdg 5,1 --dpad 30. Alignments are filtered to have at least 4 mutations with minimum PHRED estimate of 30 (quality cutoff for substitutions only) and minimum length 150nt. Co-occurrence of modifications at more than 4 positions is considered highly unlikely and assumed to be PCR jackpots. Duplicates are grouped using Hamming distance and directional network algorithm for clustering as implemented for UMIs in umi_tools. The consensus reads are then parsed for different classes of mutations using functions in ShapeMapper 2 library. Only substitution mutations are taken and pairwise mutation co-occurrence counts are recorded into a symmetric 2D matrix for each mutation spectrum, where rows and columns are positions of the nucleotide along the amplicon. For each amplicon, 2D matrices for each type of substitution mutations are summed. TMM factors are used to normalize the matrix per column and logarithm of normalized values are used. Based on mean-variance (per column) plot, minimum count threshold is selected. We chose to mask the nucleotide positions which average count less than the threshold. This results in filtering of nearly all G nucleotides, which have lower signal (lower modification/mutation rates) than other bases and different variance distribution. The diagonal of the matrix within 4nt is also masked due to extremely low co-variation. The matrix is further z-scaled per row and centered by the median of the entire matrix.
Supplementary_files_format_and_content: Text file includes the data matrix of correlated mutation counts

source_name: Transfected E14 ES cells

extract_protocol:
Error-prone PCR was performed as follows: 0.05U/uL Mutazyme II (Agilent, 200550), 1x Mutazyme II buffer, 1x SYBR, 200uM dNTP each, 300nM forward primer, 300nM reverse primer, 100pg Csde1 5’UTR fragment; 95°C 2min initial denaturation, 10 cycles of 95°C 30s, 63°C 20s, 72°C 1min. 100pg input amount was determined by initially varying the input amounts to determine the amount at which the PCR is in exponential phase (50% of signal plateau). These parameters result in a mutation rate of approximately 1 per 100nt. The error-prone PCR amplicon has homology arms for Gibson assembly into pcDNA5/FRT (ThermoFisher Scientific, V601020) plasmid with EGFP. The final construct has a flanking primer region for cDNA amplification upstream of the mutagenized 5’UTR and EGFP open reading frame downstream. NEB 10-beta E. coli cells (NEB, C3020K) are transformed by electroporation and plated over a total of 2000cm2 area to give ~10000 colonies. The plate is scraped and the mutagenesis library plasmid pool is purified. mESCs are grown, dissociated, and resuspended at 5x105 cells/mL. 3ug mutagenesis library plasmid pool DNA and 7.5uL Lipofectamine 2000 in 100uL OptiMEM (ThermoFisher Scientific, 31985062) are mixed with 2mL cells (1x106 cells) in mESC media. The cells are incubated for 10min in suspension, centrifuged, washed with mESC media, and plated into one well in a 6-well plate. 24 hours after transfection, the cells are washed, trypsinized and resuspended in 3.5mL mESC media. 1mL 1M bicine (Millipore Sigma, B3876), titrated to pH 8.5 at 25°C, is added to resuspended cells (250mM final bicine concentration). 500uL 16% dimethyl sulfate (MilliporeSigma, D186309) in ethanol is added (1.6% final concentration). Cells are mixed and incubated for 6min at 37°C. 2.5mL ice-cold 30% BME (MilliporeSigma, M3148) in ethanol is added to quench the reaction. Following centrifuge to remove the supernatant, the cells are lysed in Trizol (ThermoFisher Scientific, 15596026). Total RNA is phase extracted with chloroform and aqueous phase is purified on silica columns (Zymo, R1013). 10-20ug RNA is DNase treated at 37°C for 30min using 0.2U/uL TURBO DNase (ThermoFisher Scientific, AM2238) with 1U/uL Superase-In (ThermoFisher Scientific, AM2696) in 60uL and purified again on silica column.
1ug RNA is mixed with 1uL 1uM RT primer targeting the downstream EGFP and denatured in 6.25uL total volume (with H2O) at 65°C for 2min, then chilled to 4°C. Reverse transcription reaction conditions are as follows: 20mM Tris-HCl pH7.5, 75mM KCl, 10mM MgCl2, 5mM DTT, 500nM TGIRT (InGex, TGIRT50), 1U/uL Superase-In, 19uL total reaction volume. The RT reaction is preincubated for 25°C for 30min, then initiated with addition of 1uL 12.5mM dNTP each. After incubation at 60°C for 1hour, 1uL 2.5M NaOH is added and the reaction is heated at 95°C for 3min. 1uL 2.5M HCl and 1uL 500mM Tris-HCl pH 7.5 is added to neutralize. 29uL SPRIselect beads (Beckman Coulter, B23318) is used for purification of cDNA; elution volume is 7uL. For each of the 3 tiling primer pairs, PCR reaction is performed as follows: 0.2mM dNTP, 300nM forward primer, 300nM reverse primer, 1x SYBR Green I (ThermoFisher Scientific, S7563), Q5 Hot Start DNA polymerase (NEB, M0493S), 1x Q5 Hot Start Reaction Buffer, 1x Q5 Hot Start High GC Enhancer, 2uL cDNA, in total 20uL reaction volume. Cycling conditions are: 98°C 30s initial denaturation, 20 cycles of 98°C 10s, 64°C 10s, 72°C 30s. The amplicons are purified on silica columns and pooled. The pooled DNA is end-prepared, Illumina adaptor sequences are ligated, adaptor-ligated DNA is size selected with SPRIselect beads for 370bp, and 3 cycle barcoding PCR is performed (NEB, E7645S). 2x150bp paired end sequencing data is generated on Illumina HiSeq 4000 at Novogene.


title: iMGL_3453_scRNAseq

description: scRNAseq_Sample 1

data_processing:
10x Genomics cellranger count algorithm (v3.0.2) was used to align fastqs and count reads mapping to the genome.
Genome_build: hg19
Supplementary_files_format_and_content: tar archive containing barcode file (TSV, column indices), feature file (TSV, row indices), and barcode-feature count matrix (HDF5).

source_name: iPSC-derived microglia cells

extract_protocol:
Live iMicroglia were purified by FACS.
Libraries were constructed according to the Chromium Single Cell 3' Gene Expression Reagent Kits User Guide (v3 Chemistry) from 10x Genomics.

tag: rs76904798 genotype

value: CC

tag: differentiation protocol

value: iPSC were differentiated to microglia-like cells according to Abud et al 2017 (PMID 28426964), McQuade et al 2018 (PMID 30577865).

value: PPMI-3453 iPSC

tag: rna population

value: total RNA


title: iMGL_4101_scRNAseq

description: scRNAseq_Sample 2

value: CT

value: iPSC were differentiated to microglia-like cells

value: PPMI-4101 iPSC