title: human_CS19_brachial_rep2

city: London

first: James

last: Briscoe

address: 1 Midland Road

country: United Kingdom

institute: The Francis Crick Institute

data_processing:
Cell Ranger v3.1.0 and bcl2fastq 2.2.0 were used to quantify expression
Cell filtering to remove low quality cells from Seurat v3.2.2
SCTransform and integration of biological replicates
Genome_build: GRCh38 ensembl-release 93 provided by 10X/Cell Ranger
Supplementary_files_format_and_content: Cell Ranger output matrices

source_name: Human embryonic brachial spinal cord at CS19

extract_protocol:
Human embryonic material (4-7 weeks of gestation) was obtained from the MRC/Wellcome-Trust (grant #006237/1) funded Human Developmental Biology Resource (HDBR57, http://www.hdbr.org) with appropriate maternal written consent and approval from the London Fulham Research Ethics Committee (18/LO/0822) and the Newcastle and North Tyneside NHS Health Authority Joint Ethics Committee (08/H0906/21+5). Embryos were freshly collected on Hibernate-E medium (A12486, Gibco) supplemented with 2% B-27 (17504001 ThermoFisher Scientific) and 2.5 mL/L Glutamax (3505006, Gibco) and kept on ice until further dissection. Trunk samples of single embryos were dissected as in (Delile et al., 2019. PMID 30846445)
RNA libraries were prepared for sequencing using Chromium Single Cell 3' v3 protocol for the human and v2 for the mouse; cDNA amplification involved 12 PCR cycles

tag: developmental stage

tag: tissue

value: brachial spinal cord


title: MC68NP2

status: Public on Apr 12 2021

type: SRA

city: New York

first: Maria

last: Casanova Acebes

state: New York

address: 1470 Madison Ave

department: Oncological Sciences

country: USA

institute: Mount Sinai School of Medicine

description: YFP positive cells

library_source: transcriptomic

data_processing:
For scRNAseq GEX libraries: Read 1 is technical: Cell barcode is bp 1-16; UMI is bp 17-26. Read 2 is biological.
Gene expression libraries were aligned to the mm10 reference transcriptome and count matrices were generated using the default Cell Ranger 2.1 workflow, using the “raw” matrix output.
Genome_build: mm10
Supplementary_files_format_and_content: .tsv
Supplementary_files_format_and_content: .mtx

molecule: total RNA

organism: Mus musculus

source_name: Monocyte and macrophages from murine lungs

extract_protocol:
Single cell suspensions were obtained from lung followed by digestion with collagenase IV 0.25mg/ml (Sigma) at 37°C for 30min followed by passing through a 70mm cell strainer and RBC lysis (RBC lysis buffer, BioLegend) for 2min at room temperature.
For each mouse scRNAseq sample, 8,500 monocytes and macrophages were sorted as CD45+LINNEG (CD3/CD19/NKP46) CD11B+LY6GNEG and encapsulated using the 10X Chromium 3’ v2 chemistry kit according to manufacturer instructions. Sequencing libraries were prepared according to manufacturer instructions. QC of cDNA and final libraries was performed by CyberGreen qPCR library quantification assay (KAPA). Samples were sequenced on an Illumina Nextseq 500 using the 75-cycle kit to a depth of 100 million reads per library.

tag: cell type

value: Enriched sorted monocytes and macrophages

tag: strain

tag: group

value: naive Cx3CR1CreER-LSL-R26-YFP


title: MC68NN2

description: YFP negative cells


title: MC68TP1

description: YFP positive cells

value: tumor bearing mice (KP tumors d30)


title: human_CS19_thoracic_rep1

molecule: polyA RNA

organism: Homo sapiens

value: thoracic spinal cord


title: MC68NN1

description: YFP negative cells


title: MC68TN2

description: YFP negative cells


title: human_CS17_thoracic_rep1

source_name: Human embryonic brachial spinal cord at CS17


title: human_CS17_thoracic_rep2


title: Rapamycin, 12h, total, biological replicate 1

type: RNA

city: Rome

first: Valerio

last: Licursi

address: via degli Apuli, 4

department: c/o Dept. Biology and Biotechnologies "C. Darwin", Sapienza University of Rome

country: Italy

institute: Institute of Molecular Biology and Pathology (IBPM), National Research Council (CNR)

description: total RNA

hyb_protocol: Arrays hybridization was performed for 16 hours at 45 °C in a GeneChip Hybridization Oven 640.

scan_protocol: Each array were scanned using the Affymetrix GeneChip Scanner 3000 7G to generate the raw data (.CEL file)

data_processing:
CEL files were first processed with the Affymetrix collection of command line programs Analysis Power Tools (APT, v1.16.0) using the apt-probeset-summarize command in order to estimate the intensity at gene level with the RMA-sketch normalization. R v3.1.2 (www.r-project.org) environment and Bioconductor tRanslatome package was used to find out the probe sets that showed significant (FDR < 0.01) differential expression between experimental conditions.
probe group file: HTA-2_0.r1.pgf
meta-probeset file: HTA-2_0.na34.hg19.transcript.csv

label: biotin

source_name: human DC, rapamycin treated, transcriptome

label_protocol: Each DNA fragment is end-labelled with biotin using terminal deoxynucleotidyltransferase (TdT) (Affymetrix).

growth_protocol: Human peripheral blood mononuclear cells were isolated from buffy coats collected from healthy donors. To generate human primary DC, isolated CD14+ monocytes were cultured with 50 ng/ml GM-CSF and 1000 U/ml IL-4 (R&D Systems) for 5 days at 0.5x10^6 cells/ml in RPMI 1640 (Lonza), supplemented with 2 mM L-glutamine and 15% Fetal Bovine Serum (BioWhittaker). At day 5 the cells were tested for their differentiation status by evaluating CD1a expression (>90% CD1a+) and lack of CD14 (>95% CD14-).

extract_protocol: Total RNA was isolated from 1x10^6 infected or not infected DC, treated or not with rapamycin using Trizol (Invitrogen, Thermo Fisher Scientific) and by following manufacturer’s recommendations. Polysome-associated RNA were prepared as previously described in Livingstone et al, 2015 PMID: 26207988

treatment_protocol: Before infection and/or treatment, cell medium was replaced with RPMI without antibiotics and supplemented with 2 mM L-glutamine and 15% FBS. The deprivation of differentiating cytokines did not affect DC survival rate, which was >90%. DC cultures were then infected with virulent Mycobacterium tuberculosis (Mtb, H37Rv) at multiplicity of infection (MOI) of 1 bacterium/cell for 16 hours. Where indicated, rapamycin was added to not infected and Mtb-infected DC for 12 hours (4 hours after Mtb infection).

tag: cell line

value: primary dendrtitic cells

tag: treatment

value: rapamycin


title: human_CS17_brachial_rep1


title: human_CS19_thoracic_rep2


title: Mtb, 16h, total, biological replicate 1

description: total RNA

source_name: human DC, MTB infected, transcriptome

value: MTB infected


title: human_CS19_brachial_rep1


title: Mtb, 16h, polysome, biological replicate 1

description: polysome RNA

source_name: human DC, MTB infected, translatome


title: 5 Gy anti-NRP1 1

city: Los Angeles

first: John

middle: P

last: Chute

state: CA

address: 615 Charles E Young Drive South OHRC 545

institute: University of California, Los Angeles (UCLA)

description:
D1
Normalized_Gene_counts.txt

data_processing:
Reads per gene were quantified using STAR - 2.7.2a [3] and mm10 (Ensembl GRCm38.97). After obtaining gene counts, the counts were normalized by CPM. The Principal Component Analysis (PCA) was applied to the transcript counts. The differential gene expressions were examined by GSA. For all results of differential gene expression analysis, statistical filter were applied, which was p<0.05, FDR<0.05, and Fold Change |FC|>2.
Partek® Flow® software, version 7.0 Copyright ©; 2019 Partek Inc., St. Louis, MO, USA.
STAR: ultrafast universal RNA-seq aligner, A Dobin, CA Davis, F Schlesinger, J Drenkow, C Zaleski, S Jha, P Batut, Bioinformatics 29 (1), 15-21Fast gapped-read alignment with Bowtie 2. Langmead B, Salzberg SL. Nat Methods. (2012)

source_name: bone marrow endothelial cells

extract_protocol:
total RNA was isolated using the Qiagen microRNeasy kit
Libraries for RNA-Seq were constructed with Clontech Kit to generate strand-specific RNA-seq libraries.  The workflow consists of poly(A) RNA selection, RNA fragmentation and double-stranded cDNA generation using a mixture of random and oligo(dT) priming, followed by end repair to generate blunt ends, adaptor ligation, strand selection, and PCR amplification to produce the final libraries
Amplified libraries were quantified by Qubit dsDNA HS (High Sensitivity) Assay Kit, and quality-checked by the Agilent 4200 TapeStation System. Different index adaptors were used for multiplexing samples in one sequencing lane. Sequencing was performed with NextSeq500 High Output sequencer to produce 75 base-pair single-end reads (1 x 75 bp).

treatment_protocol: Mice were irradiated and treated daily for 2 days with with intravenous IgG control or anti-NRP1 antibody.   At 72 hours, mice were euthanized and bone marrow endothelial cells were FACs sorted for ve-cadherin+cd45- cells.

tag: radiation dose

value: 5 Gy

tag: iv treatment

value: 10 ug anti-NRP1


title: 5 Gy control 2

description:
C2
Normalized_Gene_counts.txt

value: 10 ug IgG


title: 5 Gy control 3

description:
C3
Normalized_Gene_counts.txt


title: Non-IRR anti-NRP1 3

description:
B3
Normalized_Gene_counts.txt

value: 0 Gy


title: human_CS14_thoracic_rep1

source_name: Human embryonic brachial spinal cord at CS14


title: 5 Gy anti-NRP1 3

description:
D3
Normalized_Gene_counts.txt


title: Non-IRR anti-NRP1 1

description:
B1
Normalized_Gene_counts.txt


title: human_CS14_brachial_rep1


title: human_CS12_spinalcord_rep2

source_name: Human embryonic spinal cord at CS12

value: spinal cord


title: Non-IRR control 3

description:
A3
Normalized_Gene_counts.txt


title: human_CS12_spinalcord_rep1


title: RNA-seq of CRISPRoff with non-targeting sgRNA, replicate 2 [RAB11A-GFP tagged line]

city: Dallas

first: Jin

last: Chen

state: TX

address: 5323 Harry Hines Blvd

institute: UT Southwestern Medical Center

description:
RNAseq_RAB_NT_Rep2
RNAseq_CLTA_H2B_RAB_read_counts.txt

data_processing:
For RNA-seq, linker sequences (AGATCGGAAGAGCACACGTCTGAACTC) were removed using FASTX-clipper (FASTX- Toolkit). The reads were then aligned to the human genome (GRCh37, hg19) using the STAR (Spliced Transcripts Alignment to a Reference, version 2.5) aligner against the Gencode Gene V24lift37 transcriptome annotation. Read quantification was carried out with featureCounts (Liao et al., 2014)
For the ChIP-seq, reads were aligned to the human genome (hg19) using bowtie v2.3.2 (Langmead and Salzberg, 2012) . Alignments were processed using deepTools2 bamCoverage (Ramírez et al., 2016) , normalizing reads to 1x average coverage. The resulting bigWig files were visualized on the Integrative Genomics Viewer (IGV).
For WGBS-seq, reads were first trimmed using fastp v0.21.0 (Chen et al., 2018) and then aligned to the human genome (hg38) using bwa mem v0.7.17 (Li, 2013) and methylCtools v1.0.0 (Hovestadt et al., 2014). Putative PCR duplicates were removed using sambamba v0.8.0 (Tarasov et al., 2015), and CpG methylation values were called using methylCtools. Further analyses were performed in R v4.0.2 using the bsseq v1.24.4 and DSS v2.36.0 packages.
Genome_build: hg19, hg38
Supplementary_files_format_and_content: BigWig files for ChIP-seq, counts file for RNA-seq, and methylation call files for WGBS-seq (final columns are Methylated and Unmethylated counts)

source_name: cell lysate

growth_protocol: HEK293T (female) cells were cultured in Dulbecco’s modified eagle medium (DMEM) in 10% FBS (HyClone or VWR), 100 units/mL streptomycin, 100 µg/ml penicillin, and 2 mM glutamine.

extract_protocol:
For RNA-seq, HEK293T cells that have maintained stable silencing of target genes were harvested 33 days (ITGB1 (CD29), CD81, and CD151) or 28 days (CLTA, HIST2H2BE, RAB11A) post CRISPRoff transfection. Cells were dislodged from plates with PBS, centrifuged at 500 × g for 5 min and washed again with PBS. Total RNA was extracted using Direct-zol RNA MiniPrep (Zymo R2051). For ChiP-seq, at 30 days post transfection, 10x106 cells were crosslinked with 1% formaldehyde for 10 min at room temperature and quenched with 1.25 M glycine. Crosslinked cells were washed twice with cold PBS containing 1% HaltTM protease inhibitors (Thermo Fisher Scientific) and the cell pellets were flash frozen at –80 °C until sample preparation. Cells were lysed in lysis buffer (5 mM PIPES pH 8, 85 mM KCl, 1% Igepal, 1% protease inhibitors) for 10 min on ice. Nuclei were isolated after spinning the suspension at 2000 rpm at 4 °C for 5 min. Nuclei were lysed at 4 °C for 10 min in nuclei lysis buffer (50 mM Tris pH 8, 10 mM EDTA, 1% SDS, 1% protease inhibitors). Chromatin shearing was performed at 4°C using a Diagenode Bioruptor® Pico sonication device in 1.5 ml Bioruptor® Pico Microtubes. The shearing program was optimized to obtain 200-700 bp fragments (30 seconds on, 30 seconds off for 10 cycles). The sonicated samples were centrifuged at 13,000 rpm at 4 °C for 10 min and the supernatant was collected and diluted 5-fold in IP dilution buffer (50 mM Tris pH 7.5, 150 mM NaCl, 1 mM EDTA, 1% Igepal, 0.25% deoxycholate, 1% protease inhibitors). A fraction of the input was saved and frozen (4% of total) prior to proceeding to immunoprecipitation. Immunoprecipitations were performed overnight at 4 °C using 5 µg of anti-H3K9me3 antibody (abcam ab8898). The washing steps were performed with PierceTM Protein A/G magnetic beads (Thermo Fisher Scientific) using the following protocol: once with IP wash buffer 1 (20 mM Tris pH 8, 2 mM EDTA, 50 mM KCl, 1% Triton X-100, 0.1% SDS), twice with high salt buffer (20 mM Tris pH 8, 2 mM EDTA, 500 mM NaCl, 1% Triton X-100, 0.01% SDS), once with IP wash buffer 2 (10 mM Tris pH 8, 1 mM EDTA, 0.25 lithium chloride, 1% Igepal, 1% deoxycholate), and twice with TE buffer (10 mM Tris pH 8, 1 mM EDTA). Samples were eluted in elution buffer (50 mM sodium bicarbonate, 1% SDS) at 65 °C for 1 hour and reversed crosslinked initially in 300 mM NaCl and RNase A for 1 hour at 37 °C, followed by 63 °C overnight with Proteinase K (Thermo Fisher). The DNA samples were purified using a Zymo Clean & Concentrator kit.
For RNA-seq, library preparations were carried out using TruSeq Stranded mRNA Library Preparation Kit (Illumina RS-111-2101), starting with 1000 ng total RNA. The ChIP-seq library were prepared using the NEBNext UltraTM II DNA Library Prep kit (NEB). The WGBS-seq libraries were prepared using the EZ DNA Lightning kit (Zymo, Cat. No. D5030) and the Swift Accel-NGS Methyl-Seq DNA Library Kit (Cat. No. 30024), starting from 1 ug DNA.

value: RAB11A-GFP tagged line

tag: sgrna target

value: non-targeting

tag: crispr construct

value: CRISPRoff


title: RNA-seq of CRISPRoff with RAB11A targeting sgRNA, replicate 1 [RAB11A-GFP tagged line]

description:
RNAseq_RAB_T_Rep1
RNAseq_CLTA_H2B_RAB_read_counts.txt


title: RNA-seq of CRISPRoff with RAB11A targeting sgRNA, replicate 2 [RAB11A-GFP tagged line]

description:
RNAseq_RAB_T_Rep2
RNAseq_CLTA_H2B_RAB_read_counts.txt


title: Non-IRR control 2

description:
A2
Normalized_Gene_counts.txt


title: Non-IRR control 1

description:
A1
Normalized_Gene_counts.txt


title: human_CS17_brachial_rep2


title: RNA-seq of CRISPRoff with non-targeting sgRNA, replicate 1 [RAB11A-GFP tagged line]

description:
RNAseq_RAB_NT_Rep1
RNAseq_CLTA_H2B_RAB_read_counts.txt


title: Untreated, polysome, biological replicate 1

description: polysome RNA

source_name: human DC, untreated, translatome

value: untreated


title: RNA-seq of untransfected control, replicate 2 [HIST2H2BE-GFP tagged line]

description:
RNAseq_H2B_Un_Rep2
RNAseq_CLTA_H2B_RAB_read_counts.txt

value: HIST2H2BE-GFP tagged line


title: PRO_NELFBKO_2

city: Boston

first: Karen

last: Adelman

state: MA

address: 45 Shattuck St. LHRRB-201a

department: Biological Chemistry and Molecular Pharmacology

institute: Harvard Medical School

data_processing:
library strategy: PROT-seq
PRO-seq reads were trimmed to 41bp and subsequently trimmed via cutadapt v1.14 to remove adapters: --match-read-wildcards -m 20 -q 10
PRO-seq Read 1 was aligned (single-end) using bowtie v1.2.2: -v 2 -p 6 --best --un
RNA-seq reads were quantified using kallisto v0.45.1: -b 30 --rf-stranded --genomebam --gtf --chromosomes -t 6
RNA-seq differential expression statistics were computed via tximport v1.12.3 and DESeq2 v1.24.0
Genome_build: mm10
Supplementary_files_format_and_content: bigwig; differential expression matrix

growth_protocol: Myoblasts were harvested from WT and NELF-BscKO mice, then grown in primary myoblast growth culture for 3 days.

extract_protocol:
Nuclear run-on using permeabilized cells.  Biotin-11-NTPs were incorporated into nascent transcripts during run-on and total RNA was isolated using a column based total RNA isolation kit.
PRO-seq (Precision nuclear run-on).  Biotinylated RNA from nuclear run-on was fragmented before enrichment on streptavidin magnetic beads. Illumina compatible ssRNA adapters were ligated sequentially to the RNA ends.

treatment_protocol: Myoblasts were treated with 4-hydroxytamoxifen during growth which was replenished at 24 hour intervals. This allows activation of CreERT2 for excision of floxed sites.

tag: genotype/variation

value: NELF-BscKO

value: muscle satellite cells

tag: molecule subtype

value: nascent RNA


title: 5 Gy control 1

description:
C1
Normalized_Gene_counts.txt


title: Non-IRR anti-NRP1 2

description:
B2
Normalized_Gene_counts.txt


title: RRBS_Mixture_12

first: Robin

last: Mesnage

address: Guy's Hospital, 8th Floor, Tower Wing

institute: Kings College London

library_source: genomic

data_processing:
DNA methylation calls from RRBS data were extracted with Bismark (Krueger and Andrews 2011). The output from Bismark was then imported in R and analysed with Methylkit (Akalin et al. 2012). DNA methylation calls were annotated using RefSeq gene predictions for rats (rn6 release) with the package genomation (Akalin et al. 2014). Other annotations were retrieved using the genome wide annotation for rat tool org.Rn.eg.dbR package version 3.8.2. Statistical analysis was performed with logistic regression models fitted per CpG using Methylkit functions. P-values were adjusted to Q-values using SLIM method (Wang et al. 2011).
Genome_build: rn6.0
Supplementary_files_format_and_content: count csv file

molecule: genomic DNA

organism: Rattus norvegicus

source_name: Liver

growth_protocol: Experimental groups consisted of 12 female Sprague-Dawley rats of 8 weeks of age, treated for 90 days.

extract_protocol:
DNA was extracted from rat liver using the All Prep DNA/RNA/miRNA Universal Kit (Qiagen, Hilden, Germany) using the manufacturer’s instructions for ‘Simultaneous Purification of Genomic DNA and Total RNA from Animal and Human Tissues with no alterations. Tissue weight used was ≤ 30 mg, and samples were eluted in 30μl RNase-Free Water.  DNA quantity was measured using the Qubit® 2.0 Fluorometer (Life Technologies) with the dsDNA broad range reagent, followed by quality assessment using the Agilent 2200 Tapestation and Genomic DNA screentape (Agilent Technologies). Samples displayed high molecular weight with DINs (DNA integrity score) ranging from 7.5 to 10, and average concentrations from 370 ng/μL. All samples showed a majority of high molecular weight material and all were taken forward for processing.
A total of 100ng of total DNA was diluted and processed using the Premium Reduced Representation Bisulfite Sequencing (RRBS) Kit (Diagenode, Denville, NJ, USA) as per the manufacturer’s instructions. Briefly, DNA was digested with MspI prior to end repair, adapter ligation and size selection. Products were then amplified by qPCR and pooled in equal amounts. Pooled libraries were bisulphite converted and PCR enriched following a second qPCR amplification. Libraries were quantified using the Qubit® 2.0 Fluorometer (Life Technologies) followed by quality assessment using the Agilent 2200 Tapestation and DS1000 screentape (Agilent Technologies). Pooled libraries were loaded at 1.1M with 20% standard PhiX library (Illumina, CA, USA) and sequenced to 75 base pair single end on a NextSeq 500 (Illumina, CA, USA). Data was aligned to the rat reference genome Rn6 with Bismark (Krueger and Andrews 2011).

treatment_protocol: The treatments groups were glyphosate (50 mg/kg bw/day), MON 52276 (50 mg kg bww/day of glyphosate), and a mixture of glyphosate (0.5 mg/kg bw/day), azoxystrobin (0.2 mg/kg bw/day), boscalid (0.04 mg/kg bw/day), chlorpyrifos (0.001 mg/kg bw/day), imidacloprid (0.06 mg/kg bw/day), and thiabenzadole (0.1 mg/kg bw/day), via drinking water.

value: Sprague-Dawley

value: Mixture

tag: dose

value: EU acceptable Daily Intake

value: Liver

tag: Sex

value: female


title: RRBS_Mixture_8


title: RRBS_Mixture_10


title: RRBS_Mixture_11


title: 5 Gy anti-NRP1 2

description:
D2
Normalized_Gene_counts.txt

value: 5 Gy

value: 10 ug anti-NRP1


title: RRBS_Mixture_7


title: RRBS_Mixture_6


title: RRBS_Mixture_5


title: RRBS_Mixture_4


title: PRO_WT_2

value: wild type


title: RNA-seq of CRISPRoff with HIST2H2BE targeting sgRNA, replicate 1 [HIST2H2BE-GFP tagged line]

description:
RNAseq_H2B_T_Rep1
RNAseq_CLTA_H2B_RAB_read_counts.txt


title: PRO_NELFBKO_1


title: RNA-seq of CRISPRoff with HIST2H2BE targeting sgRNA, replicate 2 [HIST2H2BE-GFP tagged line]

description:
RNAseq_H2B_T_Rep2
RNAseq_CLTA_H2B_RAB_read_counts.txt


title: PRO_WT_1


title: RNA-seq of CRISPRoff with non-targeting sgRNA, replicate 2 [HIST2H2BE-GFP tagged line]

description:
RNAseq_H2B_NT_Rep2
RNAseq_CLTA_H2B_RAB_read_counts.txt


title: RRBS_Mixture_3


title: RRBS_MON52276_12


title: KP-GFP co-cultured w/ BM monocytes #3

description: total RNA

data_processing:
Fastqs were aligned to the mm10 reference genome,
reads were dereplicated for PCR duplicates,.
Differential expression analysis were performed with the edgeR R package.
Genome_build: mm10
Supplementary_files_format_and_content: Gene counts were generated using STAR v2.5 using quantMode GeneCounts. Differential expression analysis were performed with the edgeR R package.

source_name: KP Tumor

growth_protocol: KP cells were grown in complete media (RPMI+10%FBS+1%P/S). 70% confluent cultures were detached using trypsin 0.25% and counted.

extract_protocol:
Spheroids were digested For spheroid RNA-sequencing and imaging KP-GFP cells were grown in complete media in 96-well round bottom Ultra-Low Attachment microplates (Corning – ref. 7007) to allow spheroid formation. 2500 KP-GFP cells were grown alone or mixed with 5000 TRM or 5000 BM-Mo for 7 days. 32 spheroids per conditions (KP-GFP, KP-GFP+AMs and KP-GFP+BM-Mo) were pooled and dissociated with TrypLE Express (ThermoFisher) for 20 min at 37°C and under agitation. GFP+ cells were FACS sorted (BD FACS ARIA) for RNA extraction.Sorted KP-GFP cells under the detailed conditions
For library preparation: RNA isolation from 20000-170000 FACS-sorted GFP+ KP cells was done with the Qiagen RNeasy micro kit. 1ng of RNA was synthesized into cDNA using Smart-Seq v4 Ultra Low Input RNA Kit for Sequencing (Takara Bio). Sequencing libraries were prepared using the Low Input Library Prep Kit (Takara Bio). Libraries were sequenced on an Illumina NextSeq 500 System.
Low input library

value: KP-GFP tumoral cells (spheroids)

tag: co-culture

value: with bone-marrow monocytes


title: RRBS_Mixture_1


title: KP-GFP co-cultured w/ BM monocytes #2

description: total RNA


title: RRBS_Mixture_2


title: RRBS_MON52276_11


title: RNA-seq of untransfected control, replicate 1 [HIST2H2BE-GFP tagged line]

description:
RNAseq_H2B_Un_Rep1
RNAseq_CLTA_H2B_RAB_read_counts.txt


title: RNA-seq of CRISPRoff with non-targeting sgRNA, replicate 1 [HIST2H2BE-GFP tagged line]

description:
RNAseq_H2B_NT_Rep1
RNAseq_CLTA_H2B_RAB_read_counts.txt

value: non-targeting


title: RNA-seq of untransfected control, replicate 2 [CLTA-GFP tagged line]

description:
RNAseq_CLTA_Un_Rep2
RNAseq_CLTA_H2B_RAB_read_counts.txt

value: CLTA-GFP tagged line

value: CRISPRoff


title: RRBS_Mixture_9


title: RRBS_MON52276_9


title: RNA-seq of untransfected control, replicate 1 [CLTA-GFP tagged line]

description:
RNAseq_CLTA_Un_Rep1
RNAseq_CLTA_H2B_RAB_read_counts.txt


title: RRBS_MON52276_8


title: KP-GFP co-cultured w/ BM monocytes #1

description: total RNA


title: RRBS_MON52276_6


title: KP-GFP co-cultured w/ TRMs #3

description: total RNA

value: with lung TRMs (alveolar macrophages)


title: KP-GFP co-cultured w/ TRMs #2

description: total RNA


title: RNA-seq of CRISPRoff with CLTA targeting sgRNA, replicate 2 [CLTA-GFP tagged line]

description:
RNAseq_CLTA_T_Rep2
RNAseq_CLTA_H2B_RAB_read_counts.txt

value: CLTA


title: RRBS_MON52276_7


title: RRBS_MON52276_4


title: RRBS_MON52276_3


title: KP-GFP co-cultured w/ TRMs #1

description: total RNA


title: RRBS_MON52276_5


title: RRBS_MON52276_2


title: RNA-seq of CRISPRoff with CLTA targeting sgRNA, replicate 1 [CLTA-GFP tagged line]

description:
RNAseq_CLTA_T_Rep1
RNAseq_CLTA_H2B_RAB_read_counts.txt


title: RNA-seq of CRISPRoff with non-targeting sgRNA, replicate 2 [CLTA-GFP tagged line]

description:
RNAseq_CLTA_NT_Rep2
RNAseq_CLTA_H2B_RAB_read_counts.txt


title: KP-GFP  #3

description: total RNA

value: alone


title: RNA-seq of CRISPRoff with non-targeting sgRNA, replicate 1 [CLTA-GFP tagged line]

description:
RNAseq_CLTA_NT_Rep1
RNAseq_CLTA_H2B_RAB_read_counts.txt


title: RRBS_MON52276_1


title: RRBS_Glyphosate 50_11

value: Glyphosate


title: RRBS_Glyphosate 50_10

value: Sprague-Dawley


title: KP-GFP  #1

description: total RNA


title: RRBS_Glyphosate 50_9


title: RRBS_Glyphosate 50_7


title: RNA-seq of untransfected control, replicate 1 [WT]

description:
RNAseq_Un_Rep1
RNAseq_CD29_CD81_CD151_read_counts.txt

value: WT


title: RRBS_Glyphosate 50_12


title: KP tumor d35 Ams #3

description: counts.csv [column I]

source_name: Lung tissue-resident macrophages (alveolar macrophages, Ams)

growth_protocol: KP cells were grown in complete media (RPMI+10%FBS+1%P/S). 70% confluent cultures were detached using trypsin 0.25% and counted. 5x105 KP or KP-GFP cells were intravenously injected in PBS1X into the tail vein and allowed to form tumor foci for 15 to 30 days before harvesting the lungs.

extract_protocol:
Sorted TRMs in the lung of naïve and KP-tumor bearing mice at 2 different time points (d15 and d35 post-tumor transplantation).
For library preparation: RNA isolation from 20000 FACS-sorted TRM was done with the Qiagen RNeasy micro kit. 1ng of RNA was synthesized into cDNA using Smart-Seq v4 Ultra Low Input RNA Kit for Sequencing (Takara Bio). Sequencing libraries were prepared using the Low Input Library Prep Kit (Takara Bio). Libraries were sequenced on an Illumina NextSeq 550 System.

tag: samplte type

value: d35 KP tumor


title: RRBS_Glyphosate 50_6

value: Liver


title: KP-GFP  #2

description: total RNA


title: RRBS_Glyphosate 50_5

value: female


title: RNA-seq of untransfected control, replicate 2 [WT]

description:
RNAseq_Un_Rep2
RNAseq_CD29_CD81_CD151_read_counts.txt


title: RRBS_Glyphosate 50_4


title: RRBS_Glyphosate 50_8


title: RRBS_Glyphosate 50_3


title: RRBS_Glyphosate 50_2


title: RRBS_Glyphosate 50_1


title: RNA-seq of CRISPRoff with non-targeting sgRNA, replicate 2 [WT]

description:
RNAseq_NT_Rep2
RNAseq_CD29_CD81_CD151_read_counts.txt


title: KP tumor d35 Ams #2

description: counts.csv [column H]


title: RRBS_Control_12

value: Water


title: RRBS_Control_11


title: KP tumor d15 Ams #1

description: counts.csv [column E]

value: d15 KP tumor


title: RNA-seq of CRISPRoff with non-targeting sgRNA, replicate 1 [WT]

description:
RNAseq_NT_Rep1
RNAseq_CD29_CD81_CD151_read_counts.txt


title: RNA-seq of CRISPRoff with CD81 targeting sgRNA, replicate 2 [WT]

description:
RNAseq_CD81_Rep2
RNAseq_CD29_CD81_CD151_read_counts.txt


title: KP tumor d35 Ams #1

description: counts.csv [column G]


title: RRBS_Control_10


title: Naïve Ams #3

description: counts.csv [column D]

value: Naïve


title: RRBS_Control_9


title: RNA-seq of CRISPRoff with CD29 (ITGB1) targeting sgRNA, replicate 2 [WT]

description:
RNAseq_CD29_Rep2
RNAseq_CD29_CD81_CD151_read_counts.txt


title: Naïve Ams #2

description: counts.csv [column C]


title: RNA-seq of CRISPRoff with CD29 (ITGB1) targeting sgRNA, replicate 1 [WT]

description:
RNAseq_CD29_Rep1
RNAseq_CD29_CD81_CD151_read_counts.txt


title: RRBS_Control_7


title: RRBS_Control_8


title: RRBS_Control_6


title: RNA-seq of CRISPRoff with CD151 targeting sgRNA, replicate 2 [WT]

description:
RNAseq_CD151_Rep2
RNAseq_CD29_CD81_CD151_read_counts.txt


title: Naïve Ams #1

description: counts.csv [column B]


title: RRBS_Control_3


title: RRBS_Control_4


title: RRBS_MON52276_10


title: RRBS_Control_5


title: KP tumor d30 Ams #3

data_processing:
Mouse assembly version mm10/NCBI m38 build were used for sequence alignments with bowtie version 2.3.4.1
ATAC-seq peaks were called with MACS2 2.1.1  and the parameter -q 0.0001.
Genome_build: mm10
Supplementary_files_format_and_content: File with MACS2  peak calling of common peaks between biological replicates

growth_protocol: KP cells were grown in complete media (RPMI+10%FBS+1%P/S). 70% confluent cultures were detached using trypsin 0.25% and counted. 5x10e5 KP or KP-GFP cells were intravenously injected in PBS1X into the tail vein and allowed to form tumor foci for 15 to 30 days before harvesting the lungs.

extract_protocol:
Sorted TRMs in the lung of naïve and KP-tumor bearing mice at 2 different time points (d15 and d30 post-tumor transplantation).
For library preparation: cells were lysed and incubated for 30 min at 37 °C with 50 µl of transposase reaction, which contained 25 µl of Tagment DNA buffer (Illumina), 2.5 µl of Tagment DNA enzyme (Illumina), 0.5 µl of NP-40 and 7 µl of nuclease-free water. The transposed DNA was immediately purified with the Qiagen MinElute PCR purification kit according to the manufacturer’s instructions and eluted in a 10-µl volume. PCR amplification of the transposed DNA was done with a low-cycle number protocol and with published primers.

value: lung

value: alveolar macrophage (Am)

tag: disease state

value: KP tumor

tag: time

value: d30 post-tumor transplantation


title: RNA-seq of CRISPRoff with CD151 targeting sgRNA, replicate 1 [WT]

description:
RNAseq_CD151_Rep1
RNAseq_CD29_CD81_CD151_read_counts.txt

tag: cell type

value: CRISPRoff


title: RRBS_Control_2

tag: strain

tag: tissue


title: KP tumor d15 Ams #2

description: counts.csv [column F]


title: ChIP-seq of CRISPRoff at Day 5, with sgRNA B targeting HIST2H2BE

tag: cell line

value: HIST2H2BE-GFP tagged line


title: RNA-seq of CRISPRoff with CD81 targeting sgRNA, replicate 1 [WT]

description:
RNAseq_CD81_Rep1
RNAseq_CD29_CD81_CD151_read_counts.txt


title: RRBS_Control_1

tag: treatment


title: KP tumor d30 Ams #1


title: Mixture_11

data_processing:
RNA-seq data was analysed with Salmon. This tool was used to quantify transcript abundance by mapping the reads against a reference transcriptome (Ensembl Release Rattus Norvegicus 6.0 cDNA fasta). The Salmon output was then imported in R version 3.9 using the Bioconductor package tximport
Genome_build: rn6.0
Supplementary_files_format_and_content: count csv file

extract_protocol:
RNA was extracted from rat liver using the All Prep DNA/RNA/miRNA Universal Kit (Qiagen, Hilden, Germany) using the manufacturers instructions for ‘Simultaneous Purification of Genomic DNA and Total RNA from Animal and Human Tissues’with no alterations. Tissue weight used was ≤ 30 mg, and samples were eluted in 30μl RNase-Free Water. RNA samples were quantified with the Nanodrop 8000 spectrophotometer V2.0 (ThermoScientific, USA) and integrity was checked using the Agilent 2100 Bioanalyser (Agilent Technologies, Waldbronn, Germany). All samples had RNA integrity numbers (RIN) ≥ 7.
For RNA-seq transcriptomics, 100ng of total RNA was used for each sample for library preparation. mRNA libraries were prepared using the NEBNext® Poly(A) mRNA Magnetic Isolation Module in combination with the NEBNext® Ultra™ II Directional RNA Library Prep Kit and indexed with NEBNext® Multiplex Oligos for Illumina® (96 Index Primers) (New England Biolabs, Ipswich, Massachusetts, USA). Fragmentation was carried out using incubation conditions recommended by the manufacturer for samples with a RIN ≥7 to produce RNA sizes ≥ 300bp (94°C for 10 minutes) with first strand synthesis carried out by incubation at 42oC for 50 minutes. Modification of the manufacturer’s conditions was used when enriching Adaptor Ligated DNA for libraries with a size of 300bp and 13 cycles of PCR were performed for final library enrichment. Resulting libraries were quantified using the Qubit 2.0 spectrophotometer (Life Technologies, California, USA) and average fragment size assessed using the Agilent 2200 Tapestation (Agilent Technologies, Waldbronn, Germany). Sample libraries were combined in equimolar amounts into a single pool. The final library pool was sequenced twice on the NextSeq500 at 1.1pM and 75bp paired-end reads were generated for each library using the Illumina NextSeq®500 v2.5 High-output 150 cycle kit (Illumina Inc., Cambridge, UK).

treatment_protocol: The treatments groups were glyhposate (50 mg/kg bw/day), MON 52276 (50 mg kg bww/day of glyphosate), and a mixture of glyphosate (0.5 mg/kg bw/day), azoxystrobin (0.2 mg/kg bw/day), boscalid (0.04 mg/kg bw/day), chlorpyrifos (0.001 mg/kg bw/day), imidacloprid (0.06 mg/kg bw/day), and thiabenzadole (0.1 mg/kg bw/day), via drinking water.

value: Mixture

value: EU acceptable Daily Intake


title: ChIP-seq of CRISPRoff at Day 5, with sgRNA A targeting HIST2H2BE


title: Mixture_12


title: ChIP-seq of CRISPRoff at Day 5, with non-targeting sgRNA: input DNA

value: non-targeting


title: Mixture_10

value: Sprague-Dawley


title: KP tumor d30 Ams #2


title: KP tumor d15 Ams #3

value: d15 post-tumor transplantation


title: Mixture_5


title: Mixture_6


title: KP tumor d15 Ams #2


title: KP tumor d15 Ams #4


title: ChIP-seq of CRISPRoff at Day 30, with sgRNA B targeting HIST2H2BE


title: Mixture_4


title: ChIP-seq of CRISPRoff at Day 5, with non-targeting sgRNA


title: Mixture_2


title: Mixture_3


title: Mixture_1


title: ChIP-seq of CRISPRoff at Day 30, with sgRNA A targeting HIST2H2BE


title: KP tumor d15 Ams #1


title: Naïve Ams #4

value: naïve


title: MON52276_12

value: Liver


title: ChIP-seq of CRISPRoff at Day 30, with non-targeting sgRNA


title: Naïve Ams #3


title: ChIP-seq of CRISPRoff mutant at Day 5, with sgRNA B targeting HIST2H2BE

value: CRISPRoff mutant


title: MON52276_10


title: Mixture_9


title: Naïve Ams #2


title: Mixture_7


title: MON52276_7


title: MON52276_8


title: Mixture_8


title: Naïve Ams #1


title: MON52276_6


title: ChIP-seq of CRISPRoff mutant at Day 5, with non-targeting sgRNA


title: MON52276_5


title: ChIP-seq of CRISPRoff mutant at Day 5, with sgRNA A targeting HIST2H2BE


title: AMO1 STING WT with BTZ S3

first: Mehmet

middle: Kemal

last: Samur

address: 450 Brookline Ave

institute: Dana Farber Cancer Institute

description: AMO1 cell line with BTZ treatment S3

data_processing:
For in-vitro study paired end reads were aligned to human ref genome hg19 and quantified using STAR aligner. All differential gene expression analysis were done using DESeq2.
For in-vivo study single end reads were aligned to mus musculus ref genome MM9 and quantified using STAR aligner. All differential gene expression analysis were done using DESeq2.
Genome_build: hg19 for in-vitro and MM9 for in-vivo study
Supplementary_files_format_and_content: Quantified counts with STAR in text file

source_name: multiple myeloma cell line from Bone Marrow cells

growth_protocol: For in vitro study, AMO1 WT and STING KO cell lines were cultured in in RPMI/1640 media containing 10% fetal bovine serum (FBS) 2 μmol/L glutamine, 100 U/mL penicillin, and 100 μg/mL streptomycin. For in vivo study, 5TGM1 WT and CALR KO cell lines were cultured in in Iscove's Modified Dulbecco's Media (IMDM) media supplemented with 10% FBS and 1% penicillin-streptomycin. C57BL/KaLwRijHsd mice were subcutaneously injected with 1x10 6 5TGM1 either WT or CalrKO cells in PBS.

extract_protocol: RNeasy kit from Qiagen

treatment_protocol: For in vitro study, AMO1 WT and STING KO cells were treated overnight with 5nM of BTZ. Foi in vivo study, mice were treated with BTZ that was administered intra peritoneally 0.5 mg/kg

value: AMO1 cell line


title: ChIP-seq of CRISPRoff at Day 30, with non-targeting sgRNA: input DNA


title: MON52276_3


title: AMO1 STING WT with BTZ S2

description: AMO1 cell line with BTZ treatment S2


title: MON52276_2


title: MON52276_11


title: Glyphosate 50_12

value: Glyphosate


title: ChIP-seq of CRISPRoff mutant at Day 5, with non-targeting sgRNA: input DNA


title: AMO1 STING WT S3

description: AMO1 cell line S3


title: MON52276_9


title: Glyphosate 50_11


title: Glyphosate 50_10


title: Glyphosate 50_9


title: ChIP-seq of CRISPRoff mutant at Day 30, with non-targeting sgRNA


title: Glyphosate 50_8


title: ChIP-seq of CRISPRoff mutant at Day 30, with non-targeting sgRNA: input DNA


title: AMO1 STING WT S1

description: AMO1 cell line S1


title: AMO1 STING WT S2

description: AMO1 cell line S2


title: Glyphosate 50_7


title: Glyphosate 50_6


title: RNA-seq, human, iPSC-derived 3D cerebral organoids, day 28, FXS patient3

city: Atlanta

first: Feiran

last: Zhang

state: GA

address: 615 Michael St NE

department: Human Genetics

institute: Emory University

data_processing:
For day 28 samples, paired-end reads were first aligned to human genome (hg19) by Tophat v2.1.1 with default settings. FPKM values were calculated by Cufflinks v2.2.1. Pairwise differential expression was analyzed by Cuffdiff v2.2.1.
For day 105 samples, transcript abundances were quantified using salmon 0.11.1 and an hg38 (Gencode 28) annotation. Transcript abundances were then collapsed to gene abundances using tximport 1.6.0, and differentially expressed genes were identified using DESeq2 1.18.1.
Genome_build: hg19 (day 28 samples); hg38 (day 105 samples)
Supplementary_files_format_and_content: For day 28 samples, gene expression values are provided in Cufflinks-normalized fpkm_tracking format. For day 105 samples, gene expression values are provided as TPM values as calculated by salmon.

source_name: iPSC-derived 3D cerebral organoids

growth_protocol: 3D cerebral organoids were generated from iPSC lines generated from male control and FXS patient dermal fibroblasts. Briefly, iPSCs colonies were detached from Matrigel coated plates with collagenase (1mg/ml; Invitrogen) treatment for 1hr and suspended in EB medium, comprising of FGF-2-free hESC medium supplemented with Dorsomorphin (2μM; Tocris) and A-83 (2μM; Tocris), in non-treated polystyrene plates for 4 days with a daily medium change. On days 5-6, half of the medium was replaced with induction medium consisting of DMEM/F-12, N2 Supplement (Thermofisher), 10μg/ml Heparin (Sigma), NEAA (1X), Glutamax (1X), WNT-3A (4 ng/ml; R&D Systems), CHIR99021 (1μM; Cellagentech), and SB-431542 (1μM; Cellagentech). On day 7, organoids were embedded in Matrigel (Corning) and continued to grow in induction medium for 6 more days. On day 14, embedded organoids were mechanically dissociated from Matrigel by pipetting up and down onto the plate with a 5ml pipette tip. Typically, 10 – 20 organoids were transferred to each well of a 12-well spinning bioreactor (SpinΩ) containing differentiation medium, consisting of DMEM:F12, N2 (1X), B27 (1X;  Invitrogen), 2-Mercaptoenthanol (100µM), NEAA (1X), Insulin (2.5μg/ml; Sigma). Media was changed every other day until harvested for RNA on day 28 or day 105.

extract_protocol:
For day 28 samples, total RNA was extracted from fresh 3D cerebral organoids using TRIzol (Invitrogen). Samples were treated with DNase I and cleaned with RNA Clean & Concentrator-5 kit (Zymo Research). For day 105 samples, RNA was isolated using a Zymo Quick RNA Mini kit.
For day 28 samples, mRNA-seq libraries were generated from total RNA isolated from triplicated samples per condition. The library construction was performed by Novogene. An Agilent 2100 BioAnalyzer and DNA1000 kit were used to quantify amplified cDNA and to control the quality of the libraries. For day 105 samples, mRNA-seq libraries were created using a Kapa Hyperprep mRNA kit.

tag: individual

value: FXS patient3

value: male

tag: genotype

value: FXS

value: iPSC-derived 3D cerebral organoids

tag: age

value: 28 days

tag: treatment of rna sample

value: Poly(A) selection

tag: biological replicate no.


title: Glyphosate 50_5


title: MON52276_1


title: Glyphosate 50_4


title: AMO1 STING KO with BTZ S2

description: AMO1 cell line with STING KO after BTZ treatment S2


title: Glyphosate 50_2


title: AMO1 STING WT with BTZ S1

description: AMO1 cell line with BTZ treatment S1


title: ChIP-seq of CRISPRoff mutant at Day 30, with sgRNA A targeting HIST2H2BE

value: HIST2H2BE-GFP tagged line

tag: sgrna target

tag: crispr construct


title: AMO1 STING KO with BTZ S3

description: AMO1 cell line with STING KO after BTZ treatment S3


title: RNA-seq, human, iPSC-derived 3D cerebral organoids, day 28, FXS patient1

value: FXS patient1

tag: Sex


title: AMO1 STING KO with BTZ S1

description: AMO1 cell line with STING KO after BTZ treatment S1


title: RNA-seq, human, iPSC-derived 3D cerebral organoids, day 28, control3

value: control


title: Glyphosate 50_1

tag: dose


title: AMO1 STING KO S3

description: AMO1 cell line with STING KO S3


title: RNA-seq, human, iPSC-derived 3D cerebral organoids, day 28, control2


title: Control_12

value: Water


title: Control_11

value: Sprague-Dawley


title: RNA-seq, human, iPSC-derived 3D cerebral organoids, day 28, control1


title: AMO1 STING KO S2

description: AMO1 cell line with STING KO S2


title: AMO1 STING KO S1

description: AMO1 cell line with STING KO S1


title: mouse_E85_spinalcord_rep2

data_processing:
Cell Ranger v3.1.0 and bcl2fastq 2.2.0 were used to quantify expression
Cell filtering to remove low quality cells from Seurat v3.2.2
SCTransform and integration of biological replicates
Genome_build: mm10 ensembl-release 93 provided by 10X/Cell Ranger
Supplementary_files_format_and_content: Cell Ranger output matrices

source_name: Mouse embryonic spinal cord at E8.5

extract_protocol:
Dissected samples were snipped into smaller pieces and incubated on a FACSmax (Amsbio, T200100) cell dissociation solution containing 10X Papain (30 U/mg, Sigma-Aldrich, 10108014001) for 11 min at 37°C to dissociate the cells and transferred to a HBSS solution with 5% FBS, Rock inhibitor (10 μM, Stemcell Technologies, Y-27632) and 1X of non-essential aminoacids (Thermo Fisher Scientific, 11140035). Single cells were disaggregated through pipetting and filtered, and quality control was assayed by measuring live cells versus cell death, cell size and number of clumps on an EVE cell counter. Each sample was prepared at a concentration of 600-1700 cells/µl and viability ranged between 70-96%. Samples with a viability above 70% were used for sequencing.
RNA libraries were prepared for sequencing using Chromium Single Cell 3' v2; cDNA amplification involved 12 PCR cycles

tag: developmental stage

value: spinal cord


title: Glyphosate 50_3


title: RNA-seq, human, iPSC-derived 3D cerebral organoids, day 28, FXS patient2

value: FXS patient2


title: Control_8

value: Liver


title: Control_7


title: Control_6


title: Control_5


title: macrophage null3_mock_csRNAseq

city: Irvine

first: Minji

last: Byun

state: California

address: 829 Health Science Rd, 311

department: Microbiology and Molecular Genetics

institute: University of California Irvine

data_processing:
Library strategy: capped small RNA-Seq (csRNA-seq)
Sequencing reads were trimmed for 3’ adapter sequences using HOMER and aligned using STAR with default parameters.
TSS clusters were defined using HOMER’s findcsRNATSS.pl tool that automates the discovery of strand-specific regions enriched with csRNA-seq reads found within 150 bp with a minimum read-depth of 7 reads per 107 aligned reads and >2-fold reads per bp than the surrounding 10 kb. Short RNA input libraries and total RNA-seq were used to filter out likely false-positive TSS clusters.
Genome_build: GRCh38
Supplementary_files_format_and_content: Matrix table with raw counts for every TSS cluster and every sample

source_name: hESC-derived macrophages

growth_protocol: Culture conditions for hESCs and the macrophage differentiation protocol can be found in the associated publication.

extract_protocol:
Total RNA was extracted using Zymo Quick-RNA microprep kit.
Small RNAs of ~20-60 nt were size selected from 0.4-1 µg of total RNA by denaturing gel electrophoresis. A 10% input sample was taken aside and the remainder enriched for 5’-capped RNAs with 3’-OH. Monophosphorylated RNAs were selectively degraded by Terminator 5´-Phosphate-Dependent Exonuclease (Lucigen). Subsequent 5’dephosporylation by CIP (NEB) followed by decapping with RppH (NEB) augments Cap-specific 5’adapter ligation by T4 RNA ligase 1 (NEB). The 3’ adapter was ligated using truncated T4 RNA ligase 2 (NEB) without prior 3’ repair to select against degraded RNA fragments.  Following cDNA synthesis, libraries were amplified for 14 cycles and sequenced SE75 on the Illumina NextSeq 500.

value: hESC-derived macrophages

value: untreated


title: Control_9


title: Control_4


title: macrophage null2_mock_csRNAseq


title: Control_3


title: mouse_E85_spinalcord_rep1


title: Control_2


title: macrophage null1_mock_csRNAseq


title: Control_10


title: ATAC-seq negative control

city: Seattle

first: Robert

last: Waterston

state: WA

address: Foege Building S-350D, 3720 15th Ave NE

department: Genome Sciences

institute: University of Washington

data_processing:
Convert sequencing output to fastq format (bcl2fastq v 2.19.0), split and correct barcodes, and trim adapter sequences (Trimmomatic v 0.36). These steps were performed with the sc_atac_bcl2fastq.py script.
Align reads and collapse duplicates using sc_atac_justmapping_bwa.sh script with Bowtie2 v 2.3.5 and ce11 reference.
Add a terminal letter to the cell barcode sequences (V for batch1_miseq, Y for batch2_nextseq, and Z for batch3_nextseq) to avoid collisions from different sciATAC-seq runs using modify_barcodes.py script.
Merge bam alignments with Picard MergeSamFiles v 2.15.0
Remove ce11 blacklisted regions (https://github.com/Boyle-Lab/Blacklist/blob/master/lists/ce11-blacklist.v2.bed.gz) with BedTools (v 2.25.0) and Samtools (v 1.9).
Identify the Tn5 cut sites by making a BED file containing 60 bp regions centered on the outer ends of all read pairs using the make_end_bed.py script.
Call peaks using MACS2 v 2.1.1, with genomic DNA negative control as the input.
Call valid cells based on read coverage using sc_atac_samespecies_individual_readcounter.py and sc_atac_individual_cellcall_plots.R.
Tally the peaks present in each cell in a sparse matrix file using BedTools v 2.25.0 and the peaks_to_bow2.py script.
Run iterative latent Dirichlet allocation (LDA) analysis with the lda_plus2.py script and using the sparse matrix as input. This script chooses a dimensionality for the LDA model, trains the LDA, clusters the cells based on the result, pools the data for the cells from each cluster, calls peaks for each cluster using MACS2, merges the peaks called on each cluster into a single list of genomic regions called as peaks in any cluster, and then generates a new sparse matrix that can be used in another round of LDA analysis. We conducted two rounds of LDA analysis, and we call the first round the "primary" round and the second the "refinement" round. Processed files are named as such based on from which round they were generated.
Make bigWig signal files from the MACS2 output for the refinement LDA analysis, and split multi-summit peaks into multiple contiguous peak regions using the expand_summits2.py script.
Make loom files containing the cells-by-peaks matrices and LDA results for each analysis.
Genome_build: ce11
Supplementary_files_format_and_content: *.cuts.bed:  BED file containing 60 bp regions centered on Tn5 cut sites.
*.indextable.txt:  Tab-delimited file listing the cell barcodes in the first column and metadata in the second column
*_peaks.bed:  MACS peak calling results merged into non-overlapping intervals with bedtools merge.
*.bow:  "Bag of words" file containing a sparse matrix representation of the cells-by-peaks matrix. Has three header lines: the first is the number of cells, the second is the number of peaks, and the third is not used and has arbitrary content. The rest of the file consists of three columns: The 1-based cell (row) index, the 1-based peak (column) index, and the binarized read count over that peak in that cell (always 1).
*.cell_name_to_topic_cluster_map.txt:  Tab-delimited file mapping cell barcodes to their cluster assignment from the topic-based clustering strategy.
*.loom:  File containing the the cells-by-peaks matrix annotated with metadata, including the LDA results, cluster assignments, and UMAP coordinates.
*.docTopic:  File containing the number of peaks in each cell that were assigned to each topic by the LDA generative process.
*.theta:  File containing the probability distribution over topcis for each cell based on the LDA generative process and taking into account the prior.
*.wordTopic:  File containing the number of times each peak was assigned to a particular topic by the LDA generative process.
*.phi:  File containing the probability distribution over peaks for each topic based on the LDA generative process and taking into account the prior.
*_control_lambda.bedgraph:  MACS2 output bedGraph file describing the input control signal.
*_treat_pileup.bedgraph:  MACS2 output bedGraph file describing the treatment signal.
*_peaks.narrowPeak:  MACS2 output showing peak calls in UCSC narrowPeak format.
*.summits_expanded.bed:  File containing the peak regions called by MACS2, but multi-summit peaks are split into multiple contiguous segments by detecting the trough regions between summits and splitting the peaks there.
*.subtract.bedgraph:  MACS2 output after using bdgcmp subtract to compare the treat_pileup.bedgraph and control_lambda.bedgraph files.
*.subtract.bw:  BigWig file representation of the subtract.bedgraph file.
*_primary_lda*:  Files pertaining to the first round of our iterative LDA analysis procedure.
*_refinement_lda*:  Files pertaining to the second round of our iterative LDA analysis procedure.
*_lda.filtered_peaks_iqr4.0_low_cells.*:  LDA input files after filtering out the low-coverage cells and the low- and high-prevalence peaks to make the LDA training more efficient.
*_lda.topic*:  Files pertaining to peak calling on the pooled data from the cluster of cells that have high probability in the specified topic.
*.split.trimmed.fastq.gz:  Raw fastq files with error-corrected cell barcodes attached to the read names, and Illumina sequencing adapter sequence trimmed from the ends of any short-insert reads.
all_sciATAC_merged.blacklist_filtered.bam:  Aligned reads from the *.split.trimmed.fastq.gz files, filtered for MAPQ > 10, passing QC filter, and mapping outside of the ENCODE ce11 blacklist regions.
README.txt:  Explains the relationships between the samples and the processed data files

organism: Caenorhabditis elegans

source_name: whole nematodes

growth_protocol: Worms were bleach-synchronized for at least two rounds and grown on 150 mm agar plates at 21 C at a density of about 60,000 worms per plate.

extract_protocol:
Genomic DNA isolated by phenol/chloroform extraction and ethanol precipitation from whole C. elegans worms for input to bulk ATAC-seq. This sample serves as an input control to help correct our peak calls for sequence bias in the Tn5 enzyme.
Worms were flash frozen, crushed into a powder to break the worms into pieces, and then fixed in 1.1% formaldehyde. Nuclei were extracted from the fixed pieces with several rounds of Dounce homogenization.
Single cell ATAC-seq libraries were generated using the sciATAC-seq protocol (Cusanovich, et al. 2015).
sciATAC-seq

value: unknown

tag: sample type

value: Naked genomic DNA


title: MON52276_4


title: macrophage haplo3_mock_csRNAseq


title: Control_1


title: input of Swc3-ΔZ yeast

city: Beijing

first: Zhuqiang

address: 15 Datun Road, Chaoyang District.

country: China

institute: Institute of Biophysics, CAS

data_processing:
Raw fastq read files were filtered by Trim Galore and aligned to yeast genome (sacCer3) using Bowtie2 software (v2.3.5.1).
Duplicates reads were discarded to remove potential PCR bias, and uniquely mapped reads were used for the subsequent analysis.
All ChIP samples were normalized to the same depth of 1 million unique mapped reads.
Genome_build: sacCer3
Supplementary_files_format_and_content: Bigwig files contain the reads density information normalized in CPM (counts per million) and can be viewed in genome browsers.

organism: Saccharomyces cerevisiae

source_name: Swc2-ΔZ input

extract_protocol:
Yeast cells were fixed with 1% formaldehyde for 30 min at room temperature, followed by quenching fixation with 0.125 M glycine for 5 min. Cell wall was disrupted using Ziconia beads and lysed in lysis buffer (50 mM Tris pH7.5, 1mM EDTA pH8.0, 1% Triton X-100, 150 mM NaCl, 1×EDTA-free protease inhibitor cocktail). Chromatin was sonicated to an average length of approximately 250 bp. The lysates were clarified by centrifugation, and the supernatants were incubated with protein A/G beads overnight at 4 °C to remove non-specific binding. Supernatant were collected by centrifugation and added anti-Htz1 antibody for incubation 4 h. The Antibody-Chromatin complex were pulled down with protein A Dynabeads and were washed twice with TBST buffer, twice with TE buffer, followed by incubation with TES buffer (50 mM Tris pH7.5, 10 mM EDTA pH8.0, 1% SDS) to elute the complexes. Following cross-link reversal, Rnase A and Proteinase K treatment, phenol-chloroform extraction and ethanol precipitation
DNA samples were used to prepared libraries with Kapa hyper prep kit and NEB Next multiples oligos for Illumina (NEB E7335) according to the operation instructions.

value: W1588-4C SWR1-Flag Swc2{delta}

value: SWR1-3Flag kanMX4 Swc2{delta}Z(del 1-135)::hphMX6

tag: antibody

value: none (input)


title: LAIR1KO normal lung

city: South San Francisco

first: Yasin

last: Senbabaoglu

address: 1 DNA Way

institute: Genentech

description:
BAM, matrix, and features files include data from both gene expression and antibody capture reads.
TotalSeq_SAM24386269

data_processing:
Demultiplexing of raw data was performed with the CellRanger mkfastq software (10X Genomics).
Reads obtained from demultiplexing were provided as input to ‘cellranger count’ (CellRanger v4.0.0), which aligned the reads to the mouse reference genome (mm10, 2020-A build by 10X) using STAR and collapsed to unique molecular identifier (UMI) counts.
Resulting gene-barcode matrices were filtered for background noise with 10X's cell-calling heuristic. Filtered gene-barcode matrices only included cells with at least 500 UMI counts
Subsequent data analysis was carried out in R 4.0.0 and the Seurat package (v3.2.0). RNA data were normalized with the LogNormalize option in the NormalizeData function. ADT data were normalized with centered log-ratio transformation.
HTODemux function was used with default parameters to assign sample-of-origin for each cell, and to annotate doublets and negatives (the cutoff for positive assignment was 99th percentile).
Genome_build: mm10-2020-A
Supplementary_files_format_and_content: barcode names, feature names (gene symbol or hashtag oligo), matrix of feature counts for cells. Cells with less than 500 UMI counts are excluded.

source_name: Normal lung

growth_protocol: Animals were maintained in a pathogen-free animal facility under 12hr light/12hr dark cycle with access to a standard rodent chow and water ad libitum

extract_protocol:
Chromium Single Cell 3’ Library and Gel Bead Kit V3 (10X Genomics)
N=4 single cell gene expression and N=4 antibody capture libraries

value: lung

tag: tumor

tag: library construction

value: 10x Genomics Chromium Single Cell 3' Library and Gel Bead Kit V3


title: LAIR1KO tumor-bearing lung

description:
BAM, matrix, and features files include data from both gene expression and antibody capture reads.
TotalSeq_SAM24386270

source_name: Tumor-bearing lung

tag: tissue


title: WT tumor-bearing lung

description:
BAM, matrix, and features files include data from both gene expression and antibody capture reads.
TotalSeq_SAM24386268

value: WT


title: H2A.Z ChIP in Swc2-ΔZN yeast

source_name: Swc2-ΔZN ChIP

tag: strain

value: SWR1-3Flag kanMX4 Swc2{delta}ZN(del 1-37)::hphMX6


title: input of Swc3-ΔZN yeast

source_name: Swc2-ΔZN input


title: H2A.Z ChIP in Swc2-ΔZ yeast

source_name: Swc2-ΔZ ChIP


title: macrophage haplo2_mock_csRNAseq

tag: cell type

tag: treatment


title: input of WT-Swc2 yeast

source_name: WT yeast input

tag: genotype

value: SWR1-3Flag kanMX4 Swc2::hphMX6


title: WT normal lung

description:
BAM, matrix, and features files include data from both gene expression and antibody capture reads.
TotalSeq_SAM24386267


title: macrophage haplo1_mock_csRNAseq


title: H2A.Z ChIP in WT-Swc2 yeast

source_name: WT yeast ChIP


title: D3_no_matrigel-4

city: Lausanne

first: Nicolas

last: Broguiere

address: AI 1208 Station 15

department: Sciences de la vie

country: Switzerland

institute: EPFL

data_processing:
The reads were trimmed for adapters with bcl2fastq v2.20.0
Alignment with star/2.7.0e with --quantMode GeneCounts
Genome_build: mm10
Supplementary_files_format_and_content: tab separated values containing transcript raw counts as lines for each condition as columns.

source_name: Mouse embryonic stem cells (CGR8)

growth_protocol: Mouse embryonic stem cells were aggregated in serum-free N2B27 medium, supplemented by Activin-A, Fgf2 and KSR. Mouse trophoblast stem cells were aggregated in 1:1 RPMI1640 /N2B27medium supplemented by FBS, Fgf4 and Heparin.

extract_protocol:
Trizol based RNA extraction
TruSeq stranded mRNA LT libraries were prepared according to Illumina protocol 15031047 Rev. E, starting from 300 ng of RNA, quantified by qubit DNA HS, profiled on TapeStation 4200, and sequenced on an Illumina HiSeq 4000 at a targeted depth of 36 Mreads/sample and paired-end read length of 81,8i,8i,81. The reads were trimmed for adapters with bcl2fastq v2.20.0, aligned to the mouse genome mm10 with STAR 2.7.0e, and a count matrix was assembled using the cellranger v4.0 curation of ENSEMBL annotations

treatment_protocol: TS aggregates were treated with 2% matrigel in ETS medium from 0h to 72h. In indicated samples, EPI aggregates were treated with 3% matrigel in EPI medium from 0h to 72h.

value: Differentiated mES-cells

value: Cultured in EPI medium


title: macrophage WT2_mock_csRNAseq

value: wild type


title: miRNA_seq_OE_hsa_mir217_C37UA39C_rep1

city: Stanford

first: Mark

last: Kay

address: 269 Campus Drive

institute: Stanford University

data_processing:
The sequencing data was processed by removing adapters with cutadapt (Version 1.16)
Trimmomatic (Version 0.33) was be used to filtering high-quality reads.
duplicated reads were removed with an in-house Perl script
5’ 8nt and 13nt degenerated barcode were removed for icSHAPE-MaP and RNA-seq respectively
Cleaned reads were mapped to hg38 reference by STAR(v2.7.1a).
For icSHAPE-MaP score calculation, Replicate samples were combined (with samtools merge),
Shapemapper (v2.1.4) was used to calculate final scores
Genome_build: sRNAs sequences less than ~200-nt in length were collected, such as miRNAs (from miRbase v22) (Kozomara et al., 2019), snoRNAs (from Gencode v26) (Harrow et al., 2012), snRNAs (from Gencode v26), tRNAs (from GtRNAdb v2.0) (Chan and Lowe, 2016), vault RNAs (from RefSeq v109) (O'Leary et al., 2016), Y RNAs (from RefSeq v109), and 5S rRNAs
Supplementary_files_format_and_content: Tab-seperated files. For reactivity score file, four columns represents RNA_name,position,nucleotide,score. For read count file, two columns represents RNA_name, read count. For files associated with miRNA-Seq, columns include "read ID", "Strand", "miRNA name", "start position", "end position", "sequence", "length", "mapped loci", and "raw reads per locus".

growth_protocol: Cells were maintained in DMEM/HIGH glucose with L-glutamine, sodium pyruvate (Thermo Scientific HyClone) and 10% fetal bovine serum in a humidified incubator at 37°C with 5% CO2

extract_protocol:
To enrich sRNA <200nt, after RNA extraction by Trizol LS, the total RNA was subjected to gel size selection.
For icSHAPE-MaP, The 8 µL purified RNA was ligated with a 3′ linker by incubation with the 3′ ligation mix (6 µL PEG8000, 1 µL 3’ linker (10 µM), 1 µL DTT (100 mM), 2 µL 10×ligation buffer, 1 µL T4 RNA ligase KQ (NEB), 1 µL RiboLock) at 25 °C for 2 hrs, followed by inactivation of enzyme at 65°C for 20 min. The 1.2 µL RT primer (10 µM) was added to the 3’ ligation mix and annealed to excess 3’linker by 75 °C 5 min, 37 °C 15 min, 25 °C 15 min. The 5’ ligation mix ( 3 µL PEG8000, 3 µL 10 mM ATP, 1 µL 10×ligation buffer, 0.5 µL RiboLock, 0.5 µL 5′ linker (Key Resource Table, 20 µM), 1 µL T4 RNA ligase I (NEB)) was added directly and incubated at 25 °C for 2 hrs. The reaction was purified with a RNA concentration kit (Zymo). The 9 µL mutation buffer (50 mM Tris-HCl pH 8.0, 500 µM dNTPs, 75 mM KCl, 10 mM DTT, 6 mM MnCl2, 1 µL RiboLock) was added to the 10 µL purified RNA. The reaction mix was incubated at 42 °C for 2 min. Then 1 µL SuperScript II (Thermo Fisher) was added to the reaction mix. The reverse transcription mix was incubated at 42 °C for 3 hrs. cDNA products were purified with a DNA concentration kit (Zymo).PCR were set up with 20 µL eluted cDNA and PCR reaction mix (0.5 µL P5 primer (Key Resource Table, 20 µM), 0.5 µL P3 index primer (Key Resource Table, 20 µM), 0.4 µL 25×SYBR Green, 20 µL 2×Phusion HF Master Mix (NEB)). PCR was performed in a qPCR machine (Agilent, Mx3000P) to monitor the amplification procedure and was programmed as follows: stage I: 98 °C for 1 min; stage II: 98 °C for 15 s, 65°C for 30 s, 72 °C for 45 s, with limited cycles.PCR products were purified with a DNA concentration kit (Zymo) and a further size selection (150~330 nt) on a 6% Native PAGE gel.
For sRNA-seq(40-200nt), 1 µg sRNA was then incubated with the end-repairing mix (2 µL 5×PNK buffer, 2 µL T4 polynucleotide kinase (NEB), 1 µL Fast AP (Thermo Fisher), 1µL RiboLock, 4 µL nuclease-free water) at 37°C for 1 hr. The 3′ ligation mix (6 µL PEG8000, 1 µL 3′ linker (Key Resource Table, preadenylated and blocked at its 3′ end with SpC3, 10 µM), 1 µL DTT (100 mM), 1 µL 10×RNA ligation buffer, 1 µL T4 RNA ligase I (NEB)) was added directly to the end-repairing mix. The ligation condition was 25°C for 3 hrs. The ligation product was size-selected on a 6% Urea-PAGE gel (Thermo Fisher) for ~60-200 nt. Gel purification was performed by crushing the gel and incubating in the RNA elution buffer (20 mM Tris-HCl pH 7.0, 2 mM EDTA pH 8.0, 300 mM NaAc pH 5.2, RiboLock 40 U/mL) with vigorous shaking at 37°C overnight. The eluate was collected by centrifuging through 0.45 um Spin-X columns (Thermo Fisher), followed by ethanol precipitation.   For reverse transcription, 9 µL purified 3′-ligation product was mixed with 1 µL RT primer (Key Resource Table, 10 µM). Samples were heated to 70°C for 5 min in a PCR block, cooled to 25°C by stepping down 1°C every 3 sec and held at 25°C. Afterwards, 8 µL mutation buffer (50 mM Tris-HCl pH 8.0, 500 µM dNTPs, 75 mM KCl, 10 mM DTT, 6 mM MnCl2), 1 µL RiboLock, and 1 µL SuperScript II (Thermo Fisher) were added. The reaction mix was incubated at 25°C for 3 min, and then 42°C for 3 hr. cDNA products were purified by running on 6% Urea-PAGE gel. The following steps, including circularization, PCR amplification and PCR products purification, were done as previously described.
For miRNA-Seq (~20 nt), library construction, Illumina sequencing and data analysis were conducted as previously described (Gu et al., 2012), except for the following modification. Reverse transcription (RT) was set up with RNAs ligated with adaptors, 1 µL RT primer, 1×SSIV buffer, 500 µM dNTP, 5 mM DTT, 1 U/µL RiboLock, and 1 µL SuperScript IV (Thermo Fisher) in a total volume of 20 µL. The reaction was programmed as follows: 25°C for 3 min, 42°C for 10 min, 50°C for 30 min, and 80°C for 10 min. RT products were then ethanol precipitated and dissolved in 10 µL H2O. To set up PCR reactions, 5 µL of RT products were mixed with 1×Phusion High-Fidelity PCR Master Mix with HF Buffer (NEB), 200 µM Solexa Forward and Reverse primers in a total volume of 50 µL. PCR was programmed as follows: stage I: 98°C for 30 s; stage II: 98°C for 10 s, 52°C for 30 s, 72°C for 15 s, repeating for total 22 cycles; Stage III: 72°C for 5 min. Multiplex sequencing was performed on an Illumina NextSeq 500 machine or on a MiSeq machine (1X36-bp) at the Stanford Functional Genomics Facility.

treatment_protocol: For Dicer RIP icSHAPE-MaP, Dicer KO 293T cells was transfected with plasmid encoded flag-tagged catalytic dead hDicer.Also, for sRNA-seq on WT/catalytic-dead Dicer overexpressed Dicer KO 293T cells, the Dicer KO 293T cells was transfected with plasmid encoded WT or calalytic-dead hDicer.

tag: cell line

value: Overexpression of hsa-mir-217 C37UA39C


title: miRNA_seq_OE_hsa_mir217_U20C_rep1

value: Overexpression of hsa-mir-217 U20C


title: miRNA_seq_OE_hsa_mir217_U20A_rep1

value: Overexpression of hsa-mir-217 U20A


title: D3_matrigel_4

value: Cultured in EPI medium+3% matrigel


title: D3_TS_agg_4

source_name: Mouse trophoblast stem cells

value: Differentiated mTS-cells

value: Cultured in ETS medium


title: macrophage WT1_mock_csRNAseq

value: hESC-derived macrophages

value: untreated


title: miRNA_seq_OE_hsa_mir217_U20CC38U_rep1

value: Overexpression of hsa-mir-217 U20CC38U


title: D3_no_matrigel-3


title: miRNA_seq_OE_hsa_mir217_WT_rep2

value: Overexpression of pre-hsa-mir-217 WT


title: D3_matrigel_3


title: miRNA_seq_OE_hsa_mir217_C38U_rep2

value: Overexpression of hsa-mir-217 C38U


title: K27Me3_R2_PBMC_(20201125_CNT_SW-DK_2_S2)

first: Jorja

last: Henikoff

address: 1100 Fairview AV N, A1-162

department: Basic Sciences

institute: Fred Hutchinson Cancer Research Center

description:
scCUT&Tag (10x GEM) for native nuclei from anonymous mixed peripheral blood mononuclear cell (PBMCs) donors.
Illumina NextSeq 2000

data_processing:
Genome_build: UCSC HG38
A. iCell8 system: 1. We used a custom python script to demultiplex and annotate fastq reads in the following format: Antibody_CellType_Rep_Cell#, e.g K27me3_K562_r1_54. 2. we used Bowtie2 2.3.4.3 with options --end-to-end --very-sensitive --no-mixed --no-discordant -q --phred33 -I 10 -X 700 to map the paired-end 25bp reads to HG38 of Homo sapiens genomic sequence obtained from UCSC. 3. We extracted properly paired reads from Homo sapiens alignments to generate a fragment bed files <chr> <start> <stop> <barcode> <duplicateCount>. Custom software to analyse these files is available from https://github.com/Henikoff/scCUT-Tag. B. 10X system: Cell Ranger ATAC v1.1 from 10X company with default parameters was used to do the demultiplexing, alignment to HG38 and read quantification.

source_name: CUT&TAG

value: H3K27me3, Cell Signaling Technology, Cat# 9733; Lot# 8

value: hESC


title: K27Me3_R1_PBMC_(20201125_CNT_SW-DK_1_S1)


title: macrophage WT3_mock_csRNAseq


title: H3K4me3 Hi-CHIP in USP22 knockdown (shUSP22), Thaps treated cells, replicate 2

city: Philadelphia

first: Priyankara

middle: J

last: Wickramasinghe

state: PA

address: 3601 Spruce Street

department: Bioinformatics

institute: The Wistar Institute

data_processing:
Library strategy: Hi-CHIP
Raw Hi-CHIP data was aligned using bowtie2 (v2.2.4) algorithm against hg19 genome version and processed using HiC-Pro (v2.11.3) software with default parameters. Loops across all samples were identified based on HiC-Pro output using hichipper (v0.7.7) R package (Ref3) and sloops that passed FDR<1% threshold were considered significant
Genome_build: hg19
Supplementary_files_format_and_content: CHIP-seq signal in bigwig format

source_name: HCT116 colorectal cancer cells

extract_protocol:
HCT116 cells were fixed in 1% formaldehyde for 10 minutes at room temperature, followed by quenching with 0.125 M Glycine for 5 minutes. Cells were harvested and crosslinked chromatin was extracted, MboI-digested, biotin-labeled, and ligated as described. Chromatin was sheared to 100-300 bp fragments using the Q800R2 Sonicator with circulating chiller (QSonica), for 25 minutes at 50% amplitude, 20 sec on/20 sec off. Sheared chromatin was incubated with anti-H3K4me3 (Sigma #: 07-473) for 2 hours at 4°C against indicated proteins followed by wash, elution, reverse-crosslinking, biotin purification, and Tn5 tagmentation steps.
Libraries were constructed using Nextera XT Index Kit (Illumina) and size selected using Ampure XP beads (Beckman-Coulter).

value: anti-H3K4me3 Millipore Sigma #07-473

value: USP22 knockdown (shUSP22)

tag: shRNA

value: T

tag: replicate


title: H3K4me3 Hi-CHIP in USP22 knockdown (shUSP22), Thaps treated cells, replicate 1


title: D3_TS_agg_3


title: D3_matrigel_2

value: Differentiated mES-cells


title: H3K4me3 Hi-CHIP in USP22 knockdown (shUSP22), untreated cells, replicate 2

value: u


title: D3_TS_agg_2


title: D3_no_matrigel-1

value: Cultured in EPI medium


title: H3K4me3 Hi-CHIP in USP22 wild type (shLuc), Thaps treated cells, replicate 2

value: wt

value: shLuc


title: H3K4me3 Hi-CHIP in USP22 wild type (shLuc), Thaps treated cells, replicate 1


title: D3_matrigel_1

value: Cultured in EPI medium+3% matrigel


title: D3_no_matrigel-2


title: D3_TS_agg_1

value: Differentiated mTS-cells

value: Cultured in ETS medium


title: D7_XAV_4

data_processing:
The reads were trimmed for adapters with bcl2fastq v2.20.0
Alignment with star/2.7.0e with --quantMode GeneCounts
Genome_build: mm10
Supplementary_files_format_and_content: tab separated values containing transcript raw counts as lines for each condition as columns (i.e. XAV treated or not at each timepoint)

growth_protocol: Mouse embryonic stem cells were aggregated in serum-free N2B27 medium, supplemented by Activin-A, Fgf2 and (KSR)

extract_protocol:
Trizol based RNA extraction
TruSeq stranded mRNA LT libraries were prepared according to Illumina protocol 15031047 Rev. E, starting from 300 ng of RNA, quantified by qubit DNA HS, profiled on TapeStation 4200, and sequenced on an Illumina HiSeq 4000 at a targeted depth of 36 Mreads/sample and paired-end read length of 81,8i,8i,81.
Illumina truseq stranded bulk RNAseq

treatment_protocol: EPI aggregates were treated with XAV939 from 0h to 72h.

value: XAV939 treatment between 0-72h


title: H3K4me3 Hi-CHIP in USP22 wild type (shLuc), untreated cells, replicate 2


title: D6_XAV_4


title: YR3

city: Xining

first: Delin

last: Qi

address: No. 251, Ningda Road

institute: Qinghai university

data_processing:
Reads obtained from the sequencing machines included raw reads containing adapters or low quality bases which would affect the following assembly and analysis. Thus, to get high quality clean reads, reads were further filtered according to the following rules: 1) Removing reads containing adapters; 2) Removing reads consisting of all A bases; 3) Removing reads containing more than 10% of unknown nucleotides (N); 4) Removing low quality reads containing more than 50% of low quality (Q-value≤20) bases
Short reads alignment tool Bowtie2 (2.2.8) was used for mapping reads to ribosome RNA (rRNA) database. The rRNA mapped reads were then removed. The remaining reads were further used in assembly and analysis of transcriptome.
The rRNA removed reads of each sample were then mapped to reference genome by TopHat2 (version 2.1.1), respectively. The alignment parameters were as follows: 1) Maximum read mismatch is 2 2) Disables the coverage based search for junctions 3) The distance between mate-pair reads is 50bp 4) The standard deviation for the distribution on inner distance between mate-pair reads is 80bp
After aligned with reference genome, unmapped reads (or mapped very poorly) were then re-aligned with Bowtie2, the enriched unmapped reads were split into smaller segments which were then used to find potential splice sites. The section and the section position of these short segments were predicted as well. A set of splice sites was built with initial unmapped reads by TopHat2 without relying on the known genes annotation. Not only be used for identifying expressed genes and their quantitative expression, the sequence alignment will also be helpful to find alternative splicing and new transcripts.
Genome_build: Oncorhynchus mykiss Omyk_1.0
Supplementary_files_format_and_content: tab-delimited text file includes count and FPKM values for each Sample …

organism: Oncorhynchus mykiss

source_name: dorsal skin

extract_protocol:
The samples were collected immediately, then flash-frozen in liquid nitrogen,and total RNA was extracted using Trizol reagent.
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: phenotype

value: yellow mutant

value: dorsal skin

tag: age

value: 45 day


title: ChIP-seq of CRISPRoff mutant at Day 30, with sgRNA B targeting HIST2H2BE

data_processing:
For RNA-seq, linker sequences (AGATCGGAAGAGCACACGTCTGAACTC) were removed using FASTX-clipper (FASTX- Toolkit). The reads were then aligned to the human genome (GRCh37, hg19) using the STAR (Spliced Transcripts Alignment to a Reference, version 2.5) aligner against the Gencode Gene V24lift37 transcriptome annotation. Read quantification was carried out with featureCounts (Liao et al., 2014)
For the ChIP-seq, reads were aligned to the human genome (hg19) using bowtie v2.3.2 (Langmead and Salzberg, 2012) . Alignments were processed using deepTools2 bamCoverage (Ramírez et al., 2016) , normalizing reads to 1x average coverage. The resulting bigWig files were visualized on the Integrative Genomics Viewer (IGV).
For WGBS-seq, reads were first trimmed using fastp v0.21.0 (Chen et al., 2018) and then aligned to the human genome (hg38) using bwa mem v0.7.17 (Li, 2013) and methylCtools v1.0.0 (Hovestadt et al., 2014). Putative PCR duplicates were removed using sambamba v0.8.0 (Tarasov et al., 2015), and CpG methylation values were called using methylCtools. Further analyses were performed in R v4.0.2 using the bsseq v1.24.4 and DSS v2.36.0 packages.
Genome_build: hg19, hg38
Supplementary_files_format_and_content: BigWig files for ChIP-seq, counts file for RNA-seq, and methylation call files for WGBS-seq (final columns are Methylated and Unmethylated counts)

source_name: cell lysate

extract_protocol:
For RNA-seq, HEK293T cells that have maintained stable silencing of target genes were harvested 33 days (ITGB1 (CD29), CD81, and CD151) or 28 days (CLTA, HIST2H2BE, RAB11A) post CRISPRoff transfection. Cells were dislodged from plates with PBS, centrifuged at 500 × g for 5 min and washed again with PBS. Total RNA was extracted using Direct-zol RNA MiniPrep (Zymo R2051). For ChiP-seq, at 30 days post transfection, 10x106 cells were crosslinked with 1% formaldehyde for 10 min at room temperature and quenched with 1.25 M glycine. Crosslinked cells were washed twice with cold PBS containing 1% HaltTM protease inhibitors (Thermo Fisher Scientific) and the cell pellets were flash frozen at –80 °C until sample preparation. Cells were lysed in lysis buffer (5 mM PIPES pH 8, 85 mM KCl, 1% Igepal, 1% protease inhibitors) for 10 min on ice. Nuclei were isolated after spinning the suspension at 2000 rpm at 4 °C for 5 min. Nuclei were lysed at 4 °C for 10 min in nuclei lysis buffer (50 mM Tris pH 8, 10 mM EDTA, 1% SDS, 1% protease inhibitors). Chromatin shearing was performed at 4°C using a Diagenode Bioruptor® Pico sonication device in 1.5 ml Bioruptor® Pico Microtubes. The shearing program was optimized to obtain 200-700 bp fragments (30 seconds on, 30 seconds off for 10 cycles). The sonicated samples were centrifuged at 13,000 rpm at 4 °C for 10 min and the supernatant was collected and diluted 5-fold in IP dilution buffer (50 mM Tris pH 7.5, 150 mM NaCl, 1 mM EDTA, 1% Igepal, 0.25% deoxycholate, 1% protease inhibitors). A fraction of the input was saved and frozen (4% of total) prior to proceeding to immunoprecipitation. Immunoprecipitations were performed overnight at 4 °C using 5 µg of anti-H3K9me3 antibody (abcam ab8898). The washing steps were performed with PierceTM Protein A/G magnetic beads (Thermo Fisher Scientific) using the following protocol: once with IP wash buffer 1 (20 mM Tris pH 8, 2 mM EDTA, 50 mM KCl, 1% Triton X-100, 0.1% SDS), twice with high salt buffer (20 mM Tris pH 8, 2 mM EDTA, 500 mM NaCl, 1% Triton X-100, 0.01% SDS), once with IP wash buffer 2 (10 mM Tris pH 8, 1 mM EDTA, 0.25 lithium chloride, 1% Igepal, 1% deoxycholate), and twice with TE buffer (10 mM Tris pH 8, 1 mM EDTA). Samples were eluted in elution buffer (50 mM sodium bicarbonate, 1% SDS) at 65 °C for 1 hour and reversed crosslinked initially in 300 mM NaCl and RNase A for 1 hour at 37 °C, followed by 63 °C overnight with Proteinase K (Thermo Fisher). The DNA samples were purified using a Zymo Clean & Concentrator kit.
For RNA-seq, library preparations were carried out using TruSeq Stranded mRNA Library Preparation Kit (Illumina RS-111-2101), starting with 1000 ng total RNA. The ChIP-seq library were prepared using the NEBNext UltraTM II DNA Library Prep kit (NEB). The WGBS-seq libraries were prepared using the EZ DNA Lightning kit (Zymo, Cat. No. D5030) and the Swift Accel-NGS Methyl-Seq DNA Library Kit (Cat. No. 30024), starting from 1 ug DNA.

value: HIST2H2BE-GFP tagged line

tag: sgrna target

tag: crispr construct

value: CRISPRoff mutant


title: H3K4me3 Hi-CHIP in USP22 wild type (shLuc), untreated cells, replicate 1

value: anti-H3K4me3 Millipore Sigma #07-473


title: Ileum organoids at 24hpi scRNAseq

city: Heidelberg

first: Sergio

last: Triana

address: Meyerhofstr. 1

department: Structural and Computational biology

country: Germany

institute: EMBL

data_processing:
For scRNAseq: Raw sequencing data was processed using the CellRanger software (version 3.1.0). Reads were aligned to a custom reference genome created with the reference human genome (GRCh38) and SARS-CoV-2 reference genome (NC_045512.2).
For TAP-seq: Conversion of paired end fastq files to unaligned bam using Picard Tools FastqToSam (version  2.18.14) Extraction of UMI observations using the Drop-seq tools workflow (version  1.13; http://mccarrolllab.org/dropseq/) with STAR (version 2.5.3a) for read alignment. Then Filtering for chimeric reads and UMI count extraction using an approach adapted from Dixit et al. (python version 3.6.6 ; https://www.biorxiv.org/content/10.1101/093237v1.full). Cell containing droplets were extracted using the filtered barcodes from the scRNASeq data remaining droplets were categorized as empty droplets.
Genome_build: Homo Sapiens hg38 + SARS-CoV-2 reference genome (NC_045512.2)
Supplementary_files_format_and_content: For scRNAseq: raw count matrix generated by Cell Ranger count (.H5). For TAP-seq Count tables (.counts.csv)

source_name: Ileum Organoids

growth_protocol: Stem cells containing crypts were isolated following 2 mM EDTA dissociation of tissue sample for 1 h at 4C. Crypts were spun and washed in ice cold PBS. Fractions enriched in crypts were filtered with 70 uM filters and the fractions were observed under a light microscope. Fractions containing the highest number of crypts were pooled and spun again. The supernatant was removed, and crypts were re-suspended in Matrigel. Crypts were passaged and maintained in basal and differentiation culture media

extract_protocol:
2D seeded  organoids harvested after 0 (mock), 12 and 24 hours post-infection were washed in cold PBS and incubated in TrypLE Express for 25 min at 37°C. When microscopic examination revealed that cells had reached a single cell state, they were resuspended in DMEM/F12 and spun at 500 xg for 5 mins. Supernatant was removed and the cell pellet was resuspended in PBS supplemented with 0.04% BSA and passed through a 40-μm cell strainer.
For scRNAseq - Single-cell suspensions were loaded onto the 10x Chromium controller using the 10x Genomics Single Cell 3’ Library Kit NextGem (10x Genomics) according to the manufacturer’s instructions. For TAP-seq, Targeted scRNA-seq was performed as previously described, except for using amplified cDNA from the 10X Genomics 3’ scRNA-seq protocol as input material. In short, 10 ng of amplified cDNA were used as input for the outer primer PCR and amplified with 10 PCR cycles. A second seminested PCR using 10 ng of Ampure purified outer PCR as input was performed with inner primer mix and 7 cycles of PCR. Then, a third PCR was done adding Illumina adapters

treatment_protocol: Media was removed from cells and 10^6 pfu of SARS-CoV-2 (as determined in Vero cells) was added to cells for 1 hour at 37°C. Virus was removed, cells were washed 1x with PBS and media was added back to the cells.

value: 24 hours post infection

value: Ileum Organoids


title: Ileum organoids at 24hpi Targeted TAP-seq


title: YR2


title: H3K4me3 Hi-CHIP in USP22 knockdown (shUSP22), untreated cells, replicate 1

value: USP22 knockdown (shUSP22)


title: D7_no_matrigel_4

value: Differentiated mES-cells


title: D6_no_matrigel_4


title: D5_XAV_4


title: Ileum organoids at 12hpi scRNAseq

value: 12 hours post infection


title: WR3

value: wild-type


title: D5_no_matrigel_4


title: Ileum organoids at Mock Targeted TAP-seq

value: Mock


title: Ileum organoids at 12hpi Targeted TAP-seq


title: YR1


title: WR2


title: D4_XAV_4


title: WR1


title: Ileum organoids at Mock scRNAseq


title: D4_no_matrigel_4


title: Colon organoids at 24hpi Targeted TAP-seq

source_name: Colon Organoids

value: Colon Organoids


title: D_Q040_rep2

city: Shanghai

first: Yuxuan

last: Zheng

address: 825 Zhangheng Rd.

institute: Fudan University

data_processing:
(RNA & Whole genome bisulfite sequencing) Illumina CASAVA version 1.8 were used to the basecalling.
(RNA & Whole genome bisulfite sequencing) Sequencing adapters, low quality sequences and amplification primers were trimmed from the raw sequencing reads.
(RNA) Raw RNA-seq sequencing data were firstly removed adaptors and low-quality bases, and then the clean reads were mapped to the UCSC human bosTau8 reference genome using Tophat (version: 2.0.12) with default parameters. Reads counts were calculated with HTSeq, and gene expression levels are quantified with RPKM (reads per kilobase million).
(Whole genome bisulfite sequencing) All sequencing reads were first trimmed to remove adaptors and low-quality bases using trim_galore (version 0.3.3) (parameters: --quality 20 --phred33 --length 50 --paired). Next, reads that passed quality control were mapped to the cow reference genome (bosTau8) using Bismark (version 0.7.6) with paired-end alignment mode (parameters: --non-directional --fastq). To recover additional sequencing reads, unmapped reads were realigned to the same reference genome in single-end alignment mode. Only reads with a unique mapping location in the genome were retained for further analysis.
Genome_build: bosTau8
Supplementary_files_format_and_content: "_RNA_expression.txt" files include the RPKM value and reads count of RefSeq genes. ".1X_sort_CpG.bed" files include the 'chr,start,end,base,strand,all_reads,methylated_reads,unmethylated_reads,methylation_rate,ref,type'

organism: Bos taurus

source_name: Somatic cells

extract_protocol: The WGBS libraries were constructed following the standard protocol. Briefly, genomic DNA with 0.5%unmethylated lambda DNA (Fermentas) spike-in was first sonicated into 200–300-bp fragments using Covaris S2 system. After being concentrated, the sheared DNA was end-repaired, dA-tailed and ligated with pre-methylated TruSeq DNA adapters (illumina). Bisulfite conversion was conducted with Methyl Code Bisulfite Kit (Invitrogen), after Bisulfite conversion, the converted templates were PCR amplified and quantified using Qubit ds DNA high sensitivity dye (Invitrogen). For RNA-seq library, Total RNA was extracted from 4*10^6 cells using RNeasy Mini Kit according to the manufacturer’s instructions. The mRNA was isolated from 2 μg of total RNA using the NEB Next Poly(A) mRNA Magnetic Isolation Module. Subsequently, isolated mRNA was used for RNA library construction using NEB Next ultra RNA library prep kit for Illumina. The final quality-insured libraries were sequenced on HiSeq2000 sequencer.

value: Somatic

tag: technology replication

value: biology replication #2


title: macrophage siCtrl rep1

type: genomic

hyb_protocol: Standard Illumina protocol

scan_protocol: Standard Illumina protocol

data_processing: Raw IDAT files were processed with minfi v1.32.0. Probes that have failed in one or more samples (detection pval >0.01) and cross reactive probes were removed.

label: Cy5 and Cy3

source_name: macrophage

label_protocol: Standard Illumina protocol

growth_protocol: Culture conditions for hESCs and their derivatives including macrophages can be found in the accompanying publication.

extract_protocol: Genomic DNA was extracted and purified from cells using Zymo Quick-DNA microprep kit according to standard instructions.

treatment_protocol: For the siRNA-treated samples, macrophages were transfected with indicated siRNA, washed after 24 hours, and incubated for additional 96 hours prior to genomic DNA extraction.

value: wild type

value: nontargeting siRNA

value: macrophage


title: macrophage siDNMT3A rep2

value: DNMT3A-targeting siRNA


title: macrophage siDNMT3A rep1


title: macrophage null 3

value: untreated


title: macrophage null 2


title: macrophage WT 2


title: macrophage haplo 3


title: macrophage siCtrl rep2


title: MP haplo 3

source_name: myeloid progenitor

value: myeloid progenitor


title: macrophage WT 3


title: D_iPS_rep2

source_name: Induced pluripotent stem cells

value: Induced pluripotent stem


title: D7_XAV_3

value: XAV939 treatment between 0-72h


title: macrophage haplo 2


title: D_Q040_rep1

value: biology replication #1


title: Colon organoids at 12hpi Targeted TAP-seq


title: Colon organoids at 24hpi scRNAseq

value: 24 hours post infection


title: macrophage null 1


title: MP WT 1


title: Colon organoids at 12hpi scRNAseq


title: D3_XAV_4

value: Differentiated mES-cells


title: MP WT 3


title: D3_no_matrigel_4


title: MP WT 2


title: macrophage haplo 1


title: Colon organoids at Mock Targeted TAP-seq


title: D_ES_F_rep2

source_name: Embryonic stem cells

value: Embryonic stem


title: MP null 3


title: D7_no_matrigel_3


title: MP null 1


title: D_iPS_rep1


title: MP haplo 1


title: MP null 2


title: D6_XAV_3


title: miRNA_seq_OE_hsa_mir217_C37UA39C_rep2

value: WT

value: Overexpression of hsa-mir-217 C37UA39C


title: D6_no_matrigel_3


title: D_ES_F_rep1


title: R_Q040_rep2


title: macrophage WT 1


title: D5_XAV_3


title: MP haplo 2


title: EB null 3

source_name: embryoid body

value: embryoid body


title: D5_no_matrigel_3


title: R_iPS_rep2


title: miRNA_seq_OE_hsa_mir217_U20A_rep2

value: Overexpression of hsa-mir-217 U20A


title: D4_XAV_3


title: R_Q040_rep1

value: Somatic


title: miRNA_seq_OE_hsa_mir217_U20C_rep2

value: Overexpression of hsa-mir-217 U20C


title: D4_no_matrigel_3


title: D3_XAV_3


title: R_ES_F_rep2

value: biology replication #2


title: R_ES_F_rep1


title: D3_no_matrigel_3


title: rsc2NoUV_rep2_A1_1bp

city: Pullman

first: John

middle: J.

last: Wyrick

address: Biotechnology Life Sciences 241

department: School of Molecular Biosciences

institute: Washington State University

data_processing:
Library strategy: CPD-seq
Base calling by Torrent_Suite 5.0.4
Fastq files were separated into individual libraries using A1-A6 barcodes, and the barcode sequences were trimmed, as well as the 3' nucleotide of the sequencing read, using a custom perl script
Fastq files were aligned to the yeast genome (SacCer3 or SacCer3+pUC19) using bowtie2-2.2.6 software to generate SAM files
SAM files were converted to BAM files using samtools
BAM files were converted to BED files using BEDTOOLS2 bamtoBed
Damage coordinates were mapped using custom perl script
Only reads mapping to dipyrimidine sequences (i.e., TT, TC, CT, CC), corresponding to CPD lesions were retained in BED file using custom perl script.
BED files were converted to WIG file using COUNT function of IGVTools
Background values dipyrimidine sequences were generated in WIG file using custom perl script
Genome_build: SacCer3
Supplementary_files_format_and_content: WIG files containing counts of DNA CPD lesion reads filtered for only dipyrimidine mapping sequences and including background value of 0.0 at nucleotides associated with dipyrimidines with no mapped sequencing reads. A sequencing read count was  assigned to each nucleotide in the lesion-containing dipyrimidine sequence.

source_name: Yeast cells

growth_protocol: Yeast cultures were grown in YPD (yeast extract, peptone, dextrose) to OD600=~0.8 in a yeast shaker.

extract_protocol:
Yeast genomic DNA was extracted by mixing cell pellets with 250 μl of DNA lysis buffer (2% Triton X-100, 1% SDS, 100 mM NaCl, 10 mM Tris-HCl pH 8.0, 1 mM EDTA), 250 μl of PCI (Phenol:Chloroform:Isoamyl Alcohol=25:24:1), and 150 μl of acid-washed glass beads, and vortexed vigorously twice, 2 min each time. After adding 200 μl of TE (10mM Tris-HCl, pH 7.5, 1 mM EDTA) buffer, cell lysates were centrifuged and the supernatant was mixed with 1 ml of ethanol to precipitate DNA at room temperature for 5 min. DNA pellet was dissolved in 200 μl of TE and incubated with 2 μl of RNase A (ThermoFisher Scientific) at 37˚C for 1 hr to remove RNA. DNA was subsequently purified with PCI extraction, ethanol precipitated, and disolved in sterile deionized H2O. The purified genomic DNA was quantified and 5 μg of DNA was used for each sequencing library preparation.
CPD-seq procedure is adopted from the recently published emRiboSeq method (Nature Protocols 2015 10, 1433–1444). Briefly, DNA fragments were first ligated to a double-stranded adaptor DNA trP1, and free 3'-OHs were blocked by  terminal transferase and dideoxy-ATP.  CPD lesions were  cleaved with T4 endonuclease V and apurinic/apyrimidinic (AP) endonuclease, APE 1, to generage new 3'-OHs. DNA was then ligated to adaptor  A. Up to 3 different barcoded A adaptors (A1: AAGAGGAT; A2: TTCGTGAT; A3: CCTGAGAT) were used to generate libraries for different experiments. DNA was purified with Streptavidin beads and amplified for up to 6 cycles in a PCR reaction with primers complimentary to A and trP1, and the resulting DNA libraries were used for DNA sequencing.

treatment_protocol: Cells were collected and resuspended in sterile deionized H2O and irradiated with 125 J/m2 UVC. After UV irradiation, cells were spun down and resuspended in pre-warm YPD medium (yeast extract, peptone, dextrose) for repair. Aliquots (10 ml of yeast culture) were taken before UV irradiation (No UV), immediately after UV irradiation (0hr), and at 2 hours after UV irradiation (2hr). Cells were spun down and kept at -80˚C until genomic DNA isolation.

value: rsc2 mutant

tag: repair time


title: miRNA_seq_OE_hsa_mir217_C38U_rep1

value: Overexpression of hsa-mir-217 C38U


title: miRNA_seq_OE_hsa_mir217_WT_rep1

value: Overexpression of pre-hsa-mir-217 WT


title: D7_XAV_2


title: EB haplo 3

value: untreated


title: sRNA_seq_OE_catalytic_dead_Dicer_rep4

source_name: 293T Dicer KO

value: 293T Dicer KO

value: Overexpression of catalytic-dead hDicer


title: sRNA_seq_OE_catalytic_dead_Dicer_rep3


title: miRNA_seq_OE_hsa_mir217_U20CC38U_rep2

value: Overexpression of hsa-mir-217 U20CC38U


title: D7_no_matrigel_2


title: rsc2_2hrUV_rep2_A3_1bp

value: UV irradiated 125 J/m2

value: 2 hr


title: EB null 2


title: sRNA_seq_OE_catalytic_dead_Dicer_rep2


title: Nonclassical monocytes LAIRKO5 [SAM24362401]

data_processing:
RNA-sequencing data were analyzed using HTSeqGenie (Wu et al., 2016)(Pau and Reeder, 2012) as follows:
first, reads with low nucleotide qualities (70% of bases with quality <23) or rRNA and adapter contamination were removed.
The reads that passed were then aligned to the mouse reference genome GRCm38.p5 using GSNAP (Wu and Nacu, 2010) version ‘2013-10-10-v2’ with alignment parameters: ‘-M 2 -n 10 -B 2 -i 1 -N 1 -w 200000 -E 1 --pairmax-rna=200000'.
Alignments of the reads that were reported by GSNAP as “uniquely mapping” were used for subsequent analysis.
Gene expression levels were quantified as Reads Per Kilobase of exon model per Million mapped reads normalized by size factor (nRPKM), defined as number of reads aligning to a gene in a sample / (total number of uniquely mapped reads for that sample x gene length x size factor).
Genome_build: GRCm38.p5
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

source_name: Nonclassical monocytes LAIRKO

extract_protocol:
Non-classical monocytes were sorted (purity 98%) from the bone marrow of LAIR1−/− and LAIR1+/+. 5000-10000 cells were collected and RNA was extracted using Nucleospin RNA XS (Takara) and low input RNA sequencing was performed (Clontech).
RNA libraries were prepared for sequencing using standard Illumina protocols

value: Bone marrow

value: Nonclassical monocytes

value: HiSeq Rapid SBS Kit v2


title: EB haplo 2

value: embryoid body


title: Nonclassical monocytes LAIRKO4 [SAM24362400]


title: EB null 1


title: D6_no_matrigel_2

source_name: Mouse embryonic stem cells (CGR8)

value: Differentiated mES-cells


title: rad16wtAA_2hrUV_A3_1b

description: rad16∆ WT-anchor away (AA) strain, treated with rapamycin

growth_protocol: Yeast cultures were grown in YPD (yeast extract, peptone, dextrose) to OD600=~0.8 in a yeast shaker. Anchor away strains (indicated by -AA after name) were grown to OD600 ~0.8 for mutants and ~0.4 for WT and treated with 50mg/mL rapamycin for 3 hours to conditionally deplete the tagged nuclear proteins.

extract_protocol:
Yeast genomic DNA was extracted by mixing cell pellets with 250 μl of DNA lysis buffer (2% Triton X-100, 1% SDS, 100 mM NaCl, 10 mM Tris-HCl pH 8.0, 1 mM EDTA), 250 μl of PCI (Phenol:Chloroform:Isoamyl Alcohol=25:24:1), and 150 μl of acid-washed glass beads, and vortexed vigorously twice, 2 min each time. After adding 200 μl of TE (10mM Tris-HCl, pH 7.5, 1 mM EDTA) buffer, cell lysates were centrifuged and the supernatant was mixed with 1 ml of ethanol to precipitate DNA at room temperature for 5 min. DNA pellet was dissolved in 200 μl of TE and incubated with 2 μl of RNase A (ThermoFisher Scientific) at 37˚C for 1 hr to remove RNA. DNA was subsequently purified with PCI extraction, ethanol precipitated, and disolved in sterile deionized H2O. The purified genomic DNA was quantified and 5 μg of DNA was used for each sequencing library preparation.
CPD-seq procedure is adopted from the recently published emRiboSeq method (Nature Protocols 2015 10, 1433–1444). Briefly, DNA fragments were first ligated to a double-stranded adaptor DNA trP1, and free 3'-OHs were blocked by  terminal transferase and dideoxy-ATP.  CPD lesions were  cleaved with T4 endonuclease V and apurinic/apyrimidinic (AP) endonuclease, APE 1, to generage new 3'-OHs. DNA was then ligated to adaptor  A. Up to 6 different barcoded A adaptors (A1: AAGAGGAT; A2: TTCGTGAT; A3: CCTGAGAT; A4:ATCGCGAT; A5: TACTGGAT; A6: GAACTGAT) were used to generate libraries for different experiments. DNA was purified with Streptavidin beads and amplified for up to 6 cycles in a PCR reaction with primers complimentary to A and trP1, and the resulting DNA libraries were used for DNA sequencing.

treatment_protocol: Cells were collected and resuspended in sterile deionized H2O and irradiated with 125 J/m2 UVC. After UV irradiation, cells were spun down and resuspended in pre-warm YPD medium (yeast extract, peptone, dextrose)  for non-anchor away strains and YPD + rapamycin for anchor away strains to allow for repair. Aliquots (20 ml of yeast culture) were taken before UV irradiation (No UV), immediately after UV irradiation (0hr), and at 2 hours after UV irradiation (2hr). Cells were spun down and kept at -80˚C until genomic DNA isolation.

value: rad16<delta> WT-AA


title: D5_XAV_2

value: XAV939 treatment between 0-72h


title: rad16wtAA_0hrUV_A2_1bp

value: 0 hr


title: Nonclassical monocytes LAIRKO2 [SAM24362398]


title: Colon organoids at Mock scRNAseq

value: Mock

value: Colon Organoids


title: EB WT 3

value: wild type


title: rad16sth1AA_NoUV_comb_A4_1bp

description: rad16∆ Sth1-anchor away (AA) strain, treated with rapamycin to deplete Sth1

value: rad16<delta> Sth1-AA


title: sRNA_seq_OE_WT_Dicer_rep3

value: 293T Dicer KO

value: Overexpression of WT hDicer


title: R_iPS_rep1

value: Induced pluripotent stem

value: biology replication #1


title: Nonclassical monocytes LAIRKO3 [SAM24362399]


title: sRNA_seq_OE_WT_Dicer_rep4


title: rsc2_0hrUV_rep2_A2_1bp

value: rsc2 mutant

value: UV irradiated 125 J/m2


title: D5_no_matrigel_2


title: EB haplo 1

value: untreated


title: D4_no_matrigel_2


title: Nonclassical monocytes LAIRKO1 [SAM24362397]

value: Bone marrow

value: Nonclassical monocytes

value: HiSeq Rapid SBS Kit v2


title: D4_XAV_2

value: Differentiated mES-cells


title: rad16sth1AA_2hrUV_comb_A6_1bp

value: 2 hr


title: EB WT 1

value: embryoid body


title: EB WT 2


title: Nonclassical monocytes WT5 [SAM24362396]

source_name: Nonclassical monocytes WT


title: wtAA_UV0hr_A2_1bp

description: WT-anchor away (AA) control strain, treated with rapamycin

value: WT-AA

value: 0 hr


title: sRNA_seq_OE_WT_Dicer_rep1


title: Nonclassical monocytes WT4 [SAM24362395]


title: rad16sth1AA_0hrUV_comb_A5_1bp


title: D3_XAV_2

value: XAV939 treatment between 0-72h


title: Nonclassical monocytes WT3 [SAM24362394]